LABORATORY MANUAL GENITOURINARY SYSTEM

FACULTY OF MEDICINE UNIVERSITAS PADJADJARAN 2011/2012

TABLE OF CONTENTS

Week I II III IV V

Laboratory Activity Anatomy and Embryology of The Kidney .. Histology and Pathology Anatomy of The Kidney Clinical Pathology: Urine Microscopic Examination Physiology: Renal Regulation of Fluid Balance Pharmacology: Drugs Acting on The Kidney,

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Comparison of Some Diuretic Agents VI Embryology, Anatomy, Histology, And Pathology Anatomy of Ureter, Urinary Bladder, Urethra, and External Genitalia ... VII VIII Microbiology .. Embryology, Anatomy, Histology And Pathology Anatomy of Ureter, Urinary Bladder, Urethra, and External Genitalia ...

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WEEK I

ANATOMY AND EMBRYOLOGY OF THE KIDNEY

LEARNING OBJECTIVES
After performing laboratory activity, the student should be able to: 1. Describe the anatomic topography of kidney. 2. Describe the relationship of kidney and suprarenal gland to adipose, fascial coverings, and other abdominal organs. 3. Describe the vascularization, innervations and lymphatic vessels of kidney. 4. Describe basic structure gross anatomy of kidney. 5. Describe the branches of abdominal aorta and tributaries of inferior vena cava (level of vertebrae). 6. Describe development of kidney system. 7. Describe molecular regulation of kidney development. 8. Describe abnormalities/anomalies of the kidney development. RESOURCE PERSON 1. 2. 3. 4. Fifi Veronika, dr Nani M. Yazid, drg., MKes Putri Halleyana, dr Sudradjat Sulaeman, Drs, MS

REFERENCES 1. Moore, KL and Dalley, AF. Clinical Oriented Anatomy. 5th Ed. Lippincott Williams & Wilkins. 2006. pp. 308-320. 2. Sadler, T.W. Langmans Medical Embryology. 10th Ed. Lippincott Williams and Wilkins. Philadelphia. 2006. pp. 229-256. 3. Moore, K.L., and Persaud, T.V.N. The Developing Human. Clinically Oriented Embryology. 7th Ed. Elsevier Saunders. 2003. pp. 255-285.

INTRODUCTION

Functionally, the urogenital system can be divided into two entirely different components: the urinary system and the genital system. The urinary system consists of two kidneys, two ureters, one urinary bladder, and one urethrae. The superior urinary organs (kidneys and ureters) and their vessels are primary retroperitoneal structures on the posterior abdominal wall. The kidneys do the major work of the urinary system. They produce urine that is conveyed by the ureters to the urinary bladder in the pelvis. Embryologically and anatomically, however, they are intimately interwoven. Both develop from a common mesodermal ridge (intermediate mesoderm) along the posterior wall of the abdominal cavity, and initially the excretory ducts of both systems enter a common cavity, the cloaca. Three urinary systems develop in a temporal sequence from cranial to caudal segments: The pronephros, which forms in the cervical region, is vestigial. The mesonephros, which forms in the thoracic and lumbar regions, is large and is characterized by excretory units (nephrons) and its own collecting duct, the mesonephric or wolffian duct. In the human it may function briefly, but most of the system disappears. Ducts and tubules from the mesonephros form the conduit for sperm from the testes to the urethra. In the female, these ducts regress. The metanephros, or permanent kidney, develops from two sources. It forms its own excretory tubules or nephrons like the other systems, but its collecting system originates from the ureteric bud, an outgrowth of the mesonephric duct. This bud gives rise to the ureter, renal pelvis, calyces, and the entire collecting system (Fig. 2.6). Connection between the collecting and excretory tubule systems is essential for normal development (Fig. 2.7). WT1, expressed by the mesenchyme, makes this tissue competent to respond to induction by the ureteric bud. Interactions between the bud and mesenchyme occur through production of GDNF and HGF by the mesenchyme with their tyrosine kinase receptors RET and MET, respectively, produced by the ureteric epithelium. PAX2 and WNT4, produced by the ureteric bud, cause epithelialization of the metanephric mesenchyme in preparation for excretory tubule differentiation (Fig. 2.8). Early division of the ureteric bud may lead to bifid or supernumerary kidneys with ectopic ureters (Fig. 2.9). Abnormal
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positions of the kidney, such as pelvic and horseshoe kidney, are also well known (Fig. 2.10).

PREPARATION FOR THE LABORATORY ACTIVITY: 1. Before activity, the students have to finish the homework assignment first, read the primary references, so that they will be more prepared in laboratory activity. 2. During laboratory activity, the students will be asked to explain embryologic development, to show any anatomical parts using model, interactive CD, as well as histo-pathological preparates of GUS (kidney). 3. There are pre-lab Quiz (in anatomy or embryology, or histology or anatomical pathology) before beginning the laboratory activity. 4. Dont forget to bring your: laboratory manual, atlas (anatomy, histology, etc), and laboratory coat

I. ANATOMY OF KIDNEY
A. HOMEWORK ASSIGNMENT 1. See this picture below and find what structures at no. 1-5. Explain the relationship of kidneys and suprarenal glands to adipose and fascial coverings (renal fascia, perirenal fat & pararenal fat). Also their relationship with other abdominal organs (anterior & posterior surface).

1 4 2 5 3

2. Fill the blank with correctly anatomical structures Artery Source Branches Superior suprarenal Middle suprarenal Inferior suprarenal Inferior phrenic Common iliac Median sacral Renal

Supply to

3. Fill the blank with correctly anatomical structures Vein Tributaries Drain into Right renal Left renal

Regions drained

Supra renal Inferior cava vena

4. 1 2 3 4 5 6 10 7 15 16 8 11 13

12

14

17 18 a b 20 21 22 23 24 25 26

27 28 29 30 31 32 33

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5.

See the picture below, and fill the blank

1 2 4 3

6 7

8 9 10 12 11

6. See the picture below. This is the abdominal aorta and its branches. Fill the blank with correctly answer.

....................................... 6 ....................................... 7 .... ....................................8 .......................................... 9

1 ............................................ 2 ................................................

3 ............................................................

4 ........................................................ 5 ............................................................... .............................................. 10

7. Tributaries of Inferior vena cava

.................................... 2 1 .......................................

................................ 3 4 ....................................... ................................. 7 ................................. 8 5 ....................................... 6 .......................................

9 ....................................... ................................ 10 11 ................................. 12 .................................

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8.

Innervation of the kidney 1 3 2

6 7

9. Lymphatic drainage of the kidney

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B. LABORATORY ACTIVITY

Clinical Case
During a football game between two arch rivals, the wide receiver of one team was involved in a pass pattern across the middle of the field. The quarterback was being rushed and threw the pass high. The wide receiver leapt to catch the pass and just as he did so he was "sandwiched" between the cornerback and free safety. The two defensive players hit the receiver just below the ribs on the left side, one in front and one from behind. The receiver managed to hang on to the ball but crumpled to the turf in pain. At first it was thought that he was just "shaken up," but the pain in his flank continued and became more severe. He was taken to the emergency room and examined. His vital signs were slightly elevated, but within normal range. Plain film X-rays showed no broken bones, but the margin of the left psoas major muscle was not distinguishable. Urinalysis showed blood in his urine. An IVP and CT scans were done. The IVP showed leakage of contrast into the tissue immediately around the kidney. The hemorrhage was confined to the area immediately around the kidney and extended medially toward the abdominal aorta. The diagnosis was that the kidney had been lacerated or ruptured. Immediate surgery was performed to close the laceration. The player's season ended, but he recovered uneventfully.

Questions to consider: 1. Where is the left kidney located in reference to the vertebrae, ribs, and psoas major muscle? 2. What is flank pain and why did it occur there? 3. Why was the margin of the psoas major muscle not visible? 4. How did blood get into the urine? 5. What is an IVP? 6. What confined the hemorrhage to the area around the kidney? 7. Where would be the best place to make a surgical incision to expose the kidney without going into the peritoneal cavity?

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II. EMBRYOLOGY OF KIDNEY

A. HOMEWORK ASSIGNMENT Complete the figure by naming the pointed parts:

Fig. 2.1 Transverse sections through embryos at various stages of development showing formation of nephric tubules. A. 21 days. B. 25 days. Note formation of external and internal glomeruli and the open connection between the intraembryonic cavity and the nephric tubule.

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Fig. 2.2 A. Relationship of the intermediate mesoderm of the pronephric, mesonephric, and metanephric systems. In cervical and upper thoracic regions, intermediate mesoderm is segmented; in lower thoracic, lumbar, and sacral regions it forms a solid, unsegmented mass of tissue, the nephrogenic cord. Note the longitudinal collecting duct, formed initially by the pronephros but later by the mesonephros. B. Excretory tubules of the pronephric and mesonephric systems in a 5-week embryo.

Fig. 2.3 A. Transverse section through the urogenital ridge in the lower thoracic region of a 5week embryo showing formation of an excretory tubule of the mesonephric system. Note the appearance of Bowmans capsule and the gonadal ridge. The mesonephros and gonad are 14 of the attached to the posterior abdominal wall by a broad urogenital mesentery. B. Relation gonad and the mesonephros. Note the size of the mesonephros. The mesonephric duct (wolffian duct) runs along the lateral side of the mesonephros.

Fig. 2.4 Relation of the hindgut and cloaca at the end of the fifth week. The ureteric bud penetrates the metanephric mesoderm (blastema).

Fig. 2.5 A-C Ascent of the kidney. Note the change in position between the mesonephric and metanephric systems. The mesonephric system degenerates almost entirely, and only a few remnants persist in close contact with thr gonad. In both male and female embryos, the gonads descend from their original level to a much lower position.

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B. LABORATORY ACTIVITY
1. Discuss homework materials in small group, and explain development of the kidney. 2. Explain molecular regulation of the kidney development. 3. Explain clinical correlates that appear, if any, in the development of kidney.

Complete the figure by naming the pointed parts:

Fig. 2.6 Development of the renal pelvis, calyces, and collecting tubules of the metanephros. A. 6 weeks, B. At the end of the sixth week, C. 7 weeks, D. Newborn. Note the pyramid form of the collecting tubules entering the minor calyx.

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Fig. 2.7 Development of a metanephric excretory unit. Arrows, the place where the excretory unit (blue) estblishes an open communication with the collecting system ( yellow), allowing flow of urine from the glomerulus into the collecting ducts.

Fig. 2.8 Genes involved in differentiation of the kidney. A. WT1, expressed by the mesenchyme, enables this tissue to respond to induction by the ureteric bud. GDNF and HGF, also produced by mesenchyme, interact through their receptors, RET and MET, respectively, in 17 The the ureteric bud epithelium, to stimulate growth of the bud and maintain the interactions. growth factors FGF2 and BMP7 stimulate proliferation of the mesenchyme and maintain WT1 expression. B. PAX2 and WNT4, produced by the ureteric bud, cause the mesenchyme to epithelialize in preparation for excretory tubule differentiation. Laminin and type IV collagen form a basement membrane for the epithelial cells.

Fig. 2.9 A, B. A complete and a partial double ureter. C. Possible sites of ectopic ureteral openings in the vagina, urethra, and vestibule. D, E. Photomicrographs of complete and partial duplications of the ureters ( U). Arrows, duplicated hilum; B, bladder; K, kidneys; ML, median umbilical ligament.

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Fig. 2.10 A. Unilateral pelvic kidney showing the position of the adrenal gland on the affected side. B, C. Drawing and photomicrograph, respectively, of horseshoe kidneys showing the position of the inferior mesenteric artery. BW, bladder wall; U, ureters.

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WEEK II

HISTOLOGY AND PATHOLOGY ANATOMY OF THE KIDNEY

LEARNING OBJECTIVE After performing laboratory activity, the student should be able to: 1. List the organs of the urinary system and male genital system. 2. Identify the structures and regions of a kidney and describe their organization. 3. Describe the structure, function, and location of each component of a nephron. 4. Describe the function of juxtaglomerular apparatus and identify its components 5. Trace the flow of blood through the kidney. 6. Identify the components of the glomerular filtration barrier in a diagram of a portion of a renal corpuscle. 7. Describe the pathogenesis and pathology of post streptococcal glomerulonephritis. 8. Describe the pathogenesis and pathology of chronic pyelonephritis. 9. Describe the pathogenesis and pathology of common kidney tumour. 10. Describe the pathology of polycystic kidney. RESOURCE PERSON 1. Nursiah Nasution, dr. 2. Hasrayati Agustina, dr.,SpPA 3. Hermin Aminah, dr.,SpPA REFERENCES 1. Junqueira, L.C., and Carneiro, J. Janqueiras Basic Histology. Text and Atlas. 10th Edition. Lange Medical Books M.Graw-Hill. pp 383-401 and 431-447. 2. Young, B., and Heath, J.V. Weathers Functional Histology. 4th Edition. Churchill Livingstone. pp 286-309. 3. Kumar, V., Abbas, A.K., Fausto, N. Robins and Cotran Pathologic Basis of Diseases. 7th Ed. Elsevier Saunders. pp .504-506, 960-1019.

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INTRODUCTION The urinary system includes the kidneys and the urinary tract. Kidneys are paired, beanshaped, retroperitoneal organs are located in the posterior wall of the abdominal cavity. The components of urinary tract are ureters, urinary bladder, and urethra. The lumen of the tract is characteristically lined by transitional epithelium, except for certain portions of the urethra. The kidneys filter metabolic wastes and foreign substances from the blood; regulate the ion, salt, and water concentrations of the fluids that bathe the bodys tissues; and produce rennin and erythropoietin. The collection of raw filtrate from the blood in the glomerular capillaries is only the first step in urine production. It is followed by the reabsorption of important ions, small proteins, nutrients, and much of the water. These are returned to the blood in the peritubular capillaries and vasa recta in precise proportions. The portion of the raw filtrate that is not reabsorbed constitutes the urine; it is carried by the ureters from the kidneys to the urinary bladder, where it is temporarily stored and later released through the urethra. The male genital systems consist of the external genitalia and a series of glands and ducts that produce and transport the male gametes (spermatozoa) and the seminal fluid. During laboratory activity, the students will be asked to show any histological preparations and finally also discuss about important aspects of pathogenesis and pathology. Before activity, the students have to accomplish the homework assignment first, read the primary references, so that they will be more readily performing laboratory activity.

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HOMEWORK ASSIGNMENT

Please complete these figure by naming the pointed parts!

Fig.1 The urinary system

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Fig.2 General organization of the kidney (frontal section)

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Fig.3 Basic organisation of the nephron, collecting system and renal vasculature.

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Fig.4 Cellular ultrastructure of the nephron, represented schematically. Cells of the thick ascending limb of Henles loop and the distal tubule are different in their ultra-structures and functions.

Fig.5 Renal corpuscle

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Fig.6 Schematic representation of a glomerular capillary with the visceral layer of Bowmans capsule (formed of podocytes).

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PATHOLOGY OF THE KIDNEY

1. ACUTE POST STREPTOCOCCAL GLOMERULONEPHRITIS

A 10-year-old boy had just recovered from skin infection (impetigo).Two weeks later, he developed malaise, nausea and slight fever, he passed dark brown urine.

Microscopic :

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2 3 4

1=

2=

3=

4=

5=

2. CHRONIC PYELONEPHRITIS A 40-year-old man presented to the hospital because of recurrent urinary tract infections. He complained back pain, fever, polyuria and his blood pressure was 170/100 mmHg. A renal sonogram revealed extensive scarring with pelvic and calyceal enlargement and also cortical thinning.

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Macroscopic :

Microscopic :

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3. WILMS TUMOR A 3-year-old boy presented to the hospital because of a large abdominal mass. He had hematuria, abdominal pain and constipation. USG examination revealed a large mass in his left kidney.

Macroscopic :

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Microscopic :

4.

RENAL CELL CARCINOMA

A 60-year-old man presented to the hospital because of flank pain and hematuria since 1 month ago. He also complained fever, weakness and weight loss.

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Macroscopic :

Microscopic :

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5. POLYCYSTIC KIDNEY A 5 years old girl presented to the hospital because of recurrent urinary tract infection. USG reveal cystic masses in both of her kidney

Macroscopic

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Microscopic

HOMEWORK ASSIGNMENT

A. Fill in the blank box under each the picture with microscopic and macroscopic appearance of each cases. B. Answer these questions correctly!! 1. Describe the pathogenesis of acute post streptococcal glomerulonephritis 2. Describe the pathogenesis chronic pyelonephritis. 3. Describe the pathogenesis of Wilms tumour 4. Describe the pathogenesis of renal cell carcinoma 5. Describe the pathogenesis of polycystic kidney

LABORATORY ACTIVITY
Pre-requisites: The students have to do the homework assignment and read the references as listed in the first page.
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Task: 1. Discuss the homework materials in the small group (tutorial group). 2. See the specimen under microscope and try to make a schematic draw and put the most important description based on the schematic draw.

Schematic draw

Specimen No 2:

Specimen No 3:

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Specimen No 4:

Specimen No 5:

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WEEK III CLINICAL PATHOLOGY: URINE MICROSCOPIC EXAMINATION

HOMEWORK ASSINGMENT tobe collected to your tutor at the day of lab activity

What will you do on your laboratory activity today?

I.

OBJECTIVE At the end of the activity the students will understand and can describe about: 1. The recommended of preanalyticalstage of urine examination 2. Analytical stage: The urine microscopic examination 3. Post analytical stage of urine examination 4. The interpretation of the test results 5. Factors which can affect or interfere the results

II.

REFERENCE: 1. Strassinger SK; Di Lorenzo MS; Urinalysis and Body Fluids; 4 thed; Philadelphia; F.A. Davis Company, 2001.

III.

INTRODUCTION The results of urine microscopicexamination contains information about many of the bodys major information can be obtained by inexpensive laboratory tests.

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IV.

INTENDED USE Reasons for performing the urinalysis include aiding in the diagnosis of disease, screening asymptomatic populations for undetected disorders, and monitoring the progress of disease and the effectiveness of therapy. EQUIPMENTS In this laboratory activity the students do the microscopic analysis of the urine by light microscope. Another equipments: object glass, cover glass, glove, and tissue. PROCEDURES PREPARATION/PREANALYTIC PROCEDURE 1. Urine pot: Dry, clean, leak-proof (disposable pots are recommended). 2. Midstream urine: - Patients must be provided with appropriate cleansing materials, sterile pot and instructions for cleansing and voiding. - Patients are instructed to wash their hands prior to beginning the collection. Male patients should clean the glands beginning at the urethra and withdrawing the foreskin. Female patients should separate the labia and clean the urinary meatus and surrounding area.

V.

VI.

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ANALYTICAL PROCEDURE A. PHYSICAL EXAMINATION The physical examination of urine includes the determinations of the urine color, clarity, and specific gravity. The results of the physical portion of urinalysis also can be used to confirm of or to explain findings in the microscopic examination of urinalysis.
TEST PROCEDUREOF PHYSICAL EXAMINATION Determination of color and clarity can do on urine in a clean, dry and leak-proof container.SG can also determine with a refractometer.

This procedure must be followed exactly to achieve reliable results. 1. Use your gloves 2. Collect fresh, well-mixed, uncentrifuged urine specimen in a clean dry container. Mix well immediately before using.

B. MICROSCOPIC EXAMINATION: The microscopic examination of the urineuse a sediment of urine that had centrifugated. Its purpose is to detect and to identify insoluble materials present in the urine. The blood, kidney, lower genitourinary tract, external contaminations all contribute formed elements to the urine. These include RBCs, WBCs, epithelial cells, casts, bacteria, yeast, parasites, mucus, spermatozoa, crystals, and artifacts. Examination of the urinary sediment must include both identification and quantitation of the elements present.

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Squamous epithelial cells

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Squamous epithelial cells

White blood cell and granular cast

Stained white blood cell cast

Fatty cast

Mucus

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Uric acid crystals

Calcium oxalate crystals

Triple phosphate crystals

Cystine crystals

Cholesterol crystals

POSTANALYTICAL: Recording and Reporting MICROSCOPIC EXAMINATION - RBCs: <1/hpf - WBCs: < 6/hpf - Epithelial cells: represent normal sloughing of old cell. Three types of epithel cells are seen in urine: squamous, transitional, and renal tubular. - Casts: negative
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- Bacteria, yeast, parasites: negative - Mucus: normal :a small amount. It has no clinical significance when present in either female or male urine - Spermatozoa: occasionally found in the urine of male - Crystals: depends on the pH of urine; normal in acid urine: uric acid, acid urates, and sodium urates; in alkaline urine: amorphous phosphate, triple phosphate, calcium phosphate, calcium carbonate, and ammonium biurate. In abnormal urine you can found: cystine, cholesterol, leucine, thyrocine, bilirubin, sulfonamides, ampicillin, and radiographic dye. RESULT: .. ... ... DISCUSSION: .. .. .................................. .............................................................................................................. CONCLUSION: .
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Week IV

PHYSIOLOGY: RENAL REGULATION OF FLUID BALANCE

LEARNING OBJECTIVES After performing laboratory activity, the students should be able to: 1. Describe the renal regulation of fluid balance. 2. Demonstrate and explain how the kidneys respond to the water loading and salt water loading by analyzing changes in urinary volume, specific gravity and pH. RESOURCE PERSON 1. 2. 3. 4. Dr. med. Setiawan, dr. (Physiology) Reni Farenia, dr., M.Kes (Physiology) Yuni Lazuardi, dr., M.Kes (Physiology) Nova Sylviana dr.M.Kes ( Physiology)

REFERENCES 1. Guyton AC and Hall JE. Textbook of Medical Physiology. 11 th Edition. Elsevier Saunders. pp 348-365. INTRODUCTION Human being can be fasting of foods for several days. But, this survival capacity is very limited for water. Exposing to water challenges require the capability of regulatory process which mostly involves the kidney ability. Urine volume, solute concentration, and electrolyte content are adjusted by the kidneys to maintain homeostasis of the blood. Drinking excess water or eating salty foods results in a rising blood volume, which is followed by compensatory increases in the urinary excretion of the salt and water. Diuretics are agents that increase the rate of urine formation (diuresis). These agents are ion transport inhibitors that decrease the reabsorption of Na + at different sites in the nephron. As a result, Na+ and other ions such as Cl- enter the
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urine in greater amounts rather than normal, along with water, which is carried passively to maintain osmotic equilibrium. The efficacy of the different classis of diuretics vary considerably, with the increase in secretion of Na+ varying from less than 2% for the weak, potassium-sparing diuretics, to over 20% for the potent loop diuretics. Their major clinical uses are in managing disorders involving abnormal fluid retention (edema) or in treating hypertension where their diuretic action causes decreased blood volume, which leads to reduction in blood pressure. HOMEWORK ASSIGNMENT 1. How is the mechanism of water regulation reflected by urinary excretion once a person is challenged with drinking excess water for instance in amount of 1 L? 2. What do you expect from the urinary excretion analysis (urinary volume, spesific gravity and pH) in above condition?

LABORATORY ACTIVITY Pre-requisites: The students have to do the homework assignment and read the references. Task of Renal Regulation of Fluid Balance Materials: 1. Urine collection cups 2. Urinometers 3. pH paper (pH range 39) Procedure: 1. The students void their urine into collection cups at the beginning of the laboratory session. In the analyses done in step 4, this sample will serve as thecontrol (time zero).

2. The students drink 1000 mL of water or salt containing water (4 gram of NaCl). 3. After drinking the solutions described in step 2, the students void their urine every 30 minutes for 2 hours. The urine samples are analyzed as described in step 4. 4. Each of the five urine samples collected are analyzed for volume, pH, and specific gravity, as follows: (a) Volume (mL). Measure the approximate volume of urine obtained and enter the data in the table of the laboratory report.
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(b) pH. Determine the pH of the urine samples by dipping a strip of pH paper into the urine and matching the color developed with a colorchart. The urine normally has a pH between 5.0 and 7.5. (c) Specific gravity. Determine the specific gravity of the urine samples by floating a urinometer in a cylinder (fig. 9.3) nearly filled with the specimen. Read the specific gravity at the meniscus on the urinometer scale, making sure that the urinometer float is not touching the bottom or the sides of the cylinder. The specific gravity is directly related to the amount of solutes in the urine and ranges from 1.010 to 1.025. (Pure water should have a specific gravity of 1.000.)

Worksheet Enter your data in the following table of observation: Drinking of pure water Time 0 30 60 90 120 Urine volume Urine pH Urine Specific gravity

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Drinking of salt containing water Time 0 30 60 90 120 Urine volume Urine pH Urine Specific gravity

Application of the knowledge: 1. How is the urine analysis results if a person is now challenged with the high salt containing water? 2. 3. Could you explain the regulatory mechanism involved? What would be happened in a dehydrated person?

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WEEK V

PHARMACOLOGY: DRUGS ACTING ON THE KIDNEY; COMPARISON BETWEEN SOME DIURETIC AGENTS
LEARNING OBJECTIVES After performing laboratory activity, the students should be able to: 1. Explain pharmacological properties of diuretics related to the clinical applications.

RESOURCE PERSON 1. Kuswinarti (Pharmacology) 2. Ike Husen (Pharmacology) REFERENCES 1. Benowitz, N.L. Antihypertensive agents. Katzung B.G (editor). In: Basic and Clinical Pharmacology. 10 th Edition. McGraw Hill LANGE. 2007: 159-179 2. Ives, H.E. Diuretic agents. Katzung B.G (editor). In: Basic and Clinical Pharmacology. 10 th Edition. McGraw Hill LANGE. 2007: 236-252

INTRODUCTION Diuretics are drugs that increase the excretion of Na + and water from the body by an action on the kidney. Their primary effect is to decrease the reabsorption of Na+ and Cl- from the filtrate, increase water loss being secondary to the increased excretion of NaCl. This can be achieved by a direct action on the cells of the nephron or indirectly modifying the content of the filtrate. Since a very large proportion of the NaCl and water that passes into the tubule in the glomerulus is reabsorbed, a small decrease in reabsorption can result in a marked increase in excretion.

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Before activity, the students have to accomplish the homework assignment first, read the primary references, so that they will be more ready in performing laboratory activity. MATERIALS, EQUIPMENTS AND EXPERIMENTAL SUBJECTS MATERIALS a. Diuretic Agents - HCT - Furosemide - Spironolacton b. Other a. Glycerin b. Aquadest EQUIPMENTS c. d. e. f. g. h.

Beaker glass Measuring glass Catheter Fixation board Disposable injection NGT

EXPERIMENTAL SUBJECTS Three months old healthy male rabbits with the same body weight. The rabbits are divided into 2 group: - Control group will receive aquadest only. - Treatment group will receive diuretic agents consist of HCT/Furosemid/ Spironolacton. PROCEDURES o o o o Fix the rabbit on the fixation board. Put the catheter into the ostium urethra externum. Emptying the vesica urinaria. Administer orally 15 cc of water to the first rabbit as the control and 15 cc of diuretic agent to the other (each diuretic type for different rabbit). o Collect the urine after 30 minutes and repeat it twice every 30 minutes. o Compare the volume of urine from: Control group vs treatment group Treatment group vs treatment group

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RESULT:

Group I II III IV V

Drug

Volume (ml) Volume (ml) after treatment before 120 30 minutes 60 minutes 90 minutes treatment minutes

DISCUSSION

CONCLUSION :

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HOMEWORK ASSIGNMENT: To be collected to your tutor

A. DIURETICS 1. Which is the most potent diuretic found in your experiment? Give the rational explanation of your experiment! (regarding either its mechanism of action or pharmacokinetic). 2. List the indications of furosemide! Explain the rational explanation regarding its mechanism of action! 3. Explain the side effects of furosemid. 4. Explain the indication/s of HCT and its related mechanism of action! 5. Explain the mechanism of action of spironolacton. B. Anti hypertensive agents Explain the best drug of choice and worst choice for each patient below! 1. A-50-year old well controlled type 2 diabetes, male patient recently has been diagnosed as stage II essential hypertension with normal renal function (no microalbuminemia). 2. Hypertensive patient with tachycardia and chronic open angle glaucoma. 3. Hypertensive patient with history of vasospatic angina pectoris. 4. Stage II essential hypertension patient with pregnancy. 5. Patient who tends to have exacerbated asthma. Mention antihypertensive agent should be avoided and explain the mechanism! 6. A patient has Stage III essential hypertension but normal ventricular function. After evaluating the responses to several other anti-hypertensive agents, alone and in combination, the physician places the patient on oral hydralazine. Mention antihypertensive agent/s is/are likely to be needed to manage the expected and unwanted cardio-vascular side effects of hydralazine and explain!

C. NEPHROPHARMACOLOGY 1. What are guidelines for preventing drug-induced nephrotoxicity. 2. What is drug-induced nephrotoxicity. --------ss-------51

WEEK VI

EMBRYOLOGY AND ANATOMY OF URETER, URINARY BLADDER, URETHRA, AND EXTERNAL GENITALIA
LEARNING OBJECTIVES After performing laboratory activity, the student should be able to: 1. Describe the development and abnormalities of ureter, urinary bladder, urethra and prostate. 2. Describe the anatomic topography of pelvic urinary organs. 3. Describe the anatomical aspects of ureters, urinary bladder and urethra. 4. Describe the vascularization, innervations and lymphatic vessels of ureters, urinary bladder and urethra. 5. Describe development of the indefferent embryo. 6. Describe primordium of the external genitalia. 7. Describe clinical considerations of the development of external genitalia. 8. Describe the anatomic topography of male and female genital organs. 9. Describe the anatomical aspects of male and female genital organs. 10. Describe the vascularization, innervations and lymphatic vessels of male and female genital organs.

RESOURCE PERSON 1. Sudradjat Sulaeman, Drs, MS 2. Fifi Veronika, dr 3. Nani M. Yazid, drg., MKes 4. Putri Halleyana, dr

REFERENCES 1. Moore, KL and Dalley, AF. Clinical Oriented Anatomy. 5th Ed. Lippincott Williams & Wilkins. 2006. pp. 308-320. 2. Sadler, T.W. Langmans Medical Embryology. 10th Ed. Lippincott Williams and Wilkins. Philadelphia. 2006. pp. 229-256. 3. Moore, K.L., and Persaud, T.V.N. The Developing Human. Clinically Oriented Embryology. 7th Ed. Elsevier Saunders. 2003. pp. 255-285.

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INTRODUCTION The urinary system consists of two kidneys, two ureters, one urinary bladder, and one urethra. The superior urinary organs (kidneys and ureters) and their vessels are primary retroperitoneal structures on the posterior abdominal wall. The kidneys do the major work of the urinary system. They produce urine that is conveyed by the ureters in the pelvis to the urinary bladder. The urethrae conducts urine from the bladder to the exterior. The intermediate mesoderm forms a longitudinal ridge on posterior body wall, the urogenital ridge. Part of the urogenital ridge becomes nephrogenic cord, which gives rise to the urinary system. The nephrogenic cord develops into 3 nephric structures: the pronephros, mesonephros, and metanephros. The metanephros is the caudalmost nephric stucture. It begins to form at week 5 and is functional in the fetus at about week 10 of development. The metanephros develops from an outgrowth of the mesonephric duct, the ureteric bud, and from a condensation of mesoderm within the nephrogenic cord, the metanephric mesoderm (metanephric blastema). The ureteric bud initially penetrates the metanephric blastema and then undergoes repeated divisions to form the ureters, renal pelvis, major calyces, minor calyces, and collecting tubules. The urinary bladder is formed from the upper end of the urogenital sinus, which is continuous with the allantois. The allantois becomes a fibrous cord, the urachus (median umbilical ligament in the adult). The lower ends of the mesonephric ducts become incorporated into the posterior wall of the bladder at the trigone. The mesonephric ducts eventually open into the urogenital sinus below the bladder. The transitional epithelium lining the urinary bladder is derived from endoderm because of its etiology from the urogenital sinus and gut tube. The transitional epithelium lining the ureters, renal pelvis, and major and minor calyces is derived from mesoderm because of its etiology from the ureteric bud. The female urethra is formed from the lower end of the urogenital sinus, develops as endodermal outgrowths into the surrounding mesoderm to form the urethral glands and paraurethral glands, ends at the vestibule of vagina, which also forms from the urogenital sinus. The vestibule of vagina develops endoderm outgrowths into the surrounding mesoderm to form the greater vestibular glands. The transitional epithelium and stratified squamous epithelium lining the female urethra are derived from endoderm. The male urethra: Prostatic urethra, membranous urethra, and proximal urethra. These parts of the urethra are formed from lower end of urogenital sinus. The transitional epithelium and stratified squamous epithelium lining these parts of the urethra are derived from endoderm. The prostatic urethra develops endoderm outgrowth into the surrounding mesoderm to form the prostate gland. The membranous urethra develops endoderm outgrowth into the surrounding mesoderm to form the bulbourethral glands. The proximal part of penile urethra develops endoderm outgrowth into the surrounding mesoderm to form Littres glands. The distal part of penile urethra is formed from an ingrowth of surface
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ectoderm called the glandular plate. The glandular plate joins the membranous urethra and becomes canalized to form the navicular fossa. Ectodermal septa appear lateral to the navicular fossa and become canalized to form the foreskin. The stratified squamous epithelium lining this part of the urethra is derived from ectoderm. Although the genotype of the embryo (46,XX or 46,XY) is established at fertilization, the embryo during weeks 1-6 remains in a sexually indifferent or undifferentiated stage; that is, genetically female embryos and genetically male embryos are phenotypically indistinguisable. Once the cloaca has been partitioned, the development of external genitalia can begin. This begins to occur at approximately four weeks. Early in the fourth week the mesenchyme surrounding the cloacal membrane begins to proliferate. It is critical that this mesenchyme reach a critical cell mass or only rudimentary structures are formed. The proliferation of mesenchyme is known as the genital tubercles. Development of the male external genitalia is dependent upon dihydrotestosterone which is produced by the testes. As the genital tubercle is elongating and growing to form the penis, the urogenital folds which lie on either side of the urogenital membrane begin to move towards each other forming a groove, this is known as the urethral groove. The urogenital folds fuse together on the ventral side of the developing penis, enclosing what will now become the spongy urethra. If the urogenital folds fail to close, hypospadias results.

PREPARATION FOR THE LABORATORY ACTIVITY: 1. Before activity, the students have to accomplish the homework assignment first, read the primary references, so that they will be more readily performing laboratory activity. 2. During laboratory activity, the students will be asked to explain embryologic development, to show any anatomical parts using model, interactive CD, as well as histo-pathological preparates of Urinary System. 3. There are pre-lab Quiz (in anatomy or embryology, or histology or anatomical pathology) before beginning the laboratory activity. 4. Dont forget to bring your: laboratory manual, atlas (anatomy, histology, etc), and dont forget to use the laboratory coat.

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C. HOMEWORK ASSIGNMENT I. EMBRYOLOGY

Please label the following pointed parts of the picture:

................................. ................................. 3 nephric structures: ................................. .................................

.................................

.................................

................................. ................................. .................................

Fig. 1.1 Frontal view of an embryo

................................. .................................

................................. ................................. ................................. ................................. .................................

Fig. 1.2 Lateral view of a fetal kidney.

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....................... ....................... ....................... ....................... ....................... ....................... ....................... ....................... .......................

.......................

.......................

Fig. 1-3 Lateral views depicting the formation of the bladder from the upper part of the urogenital sinus at week 4 (A), and week 7 (B).

...........................

........................... ...........................

........................... ...........................

Fig. 1.4 Diagram depicting the female urethra.

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B ..........................

.......................... ..........................

.......................... ..........................

..........................

.......................... .......................... .......................... .......................... ..........................

Fig. 1-5 (A) Diagram depicting the male urethra. (B) Enlarged view of box in A

Fig. 1.1 A,B. Indifferent stages of the external genitalia.

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Fig. 1.2 A. Development of external genitalia in the male at 10 weeks. Note the deep urethral groove flanked by the urethral folds. B. Transverse sections through the phallus during formation of the penile urethra. The urogenital groove is bridged by the urethral folds. C. Development of the glandular portion of the penile urethra. D. Newborn.

Fig. 1.3 Development of the external genitalia in the female at 5 months (A) and in the newborn (B).

Think first and answer the question correctly and concisely! 1. Describe the development of ureter, urinary bladder, and urethra (prostat). 2. Describe the abnormalities of ureter, urinary bladder, and urethra (prostat). 3. Describe clinical considerations of the development of external genitalia.
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4. Describe development of the indefferent embryo. 5. Describe primordium of the external genitalia. ------ss-----II. ANATOMY

1. Please label the following pointed parts of the picture:

2. Please label the following pointed parts of the picture:

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3. Please label the following pointed parts of the picture:

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Think first and answer the question correctly and concisely! 1. Describe the anatomical route of ureter and its relationships with other pelvic organs in male and female. 2. Mention three sites at which the ureter is normally relatively constricted. 3. Describe the anatomical characteristic of male and female urinary bladder. 4. Describe the anatomical characteristic of male and female urethrae. 5. Please explain about the vascularization, innervations and lymphatic vessels of ureters, urinary bladder and urethrae. 6. Please draw systematically the innervation of male and female genitalia. 7. Please distinguish between male and female pelvic floor by drawing it systematically. ------ss------

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D. LABORATORY ACTIVITY Pre-requisites: The students have to do the homework assignment and read the references as listed in the first page. See the specimen under microscope and try to make a schematic draw and put the most important description based on the schematic draw, or learn the worksheet below and explain the aim of each figure. EMBRYOLOGY

I.

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1. Embryology of the external genitalia a. Discus the homework materials b. Explain the pictures

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II.

ANATOMY

1. Discuss the homework materials in the small group (tutorial group). 2. Please label the following pointed parts of the picture:

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3. Please label the following pointed parts of the picture:

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4. Please fill in the following pointed numbers:

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5. Please label the following pointed parts of the picture :

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6. Please fill in the following pointed numbers:

Anatomy of the genital organs a. Using model discuss about the anatomical aspect as needed and its clinical orientation

WORKSHEET OF ANATOMY Please label the following pointed parts of the picture:

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Week VII

MICROBIOLOGY
Topic : Laboratory examination of organisms associated with urinary tract and sexual transmitted infections I. General objective After finishing this activity, the student will be able to: Understood microbiological examinations to confirm the diagnosis of urinary tract and sexual transmitted infections II. Specific objective At the end of laboratory practice, the student could: 1. Understood methods of specimen collection and the laboratory examinations to confirm the diagnosis of urinary tract and sexual transmitted infections 2. Understood the methods of examination, isolation, and identification of bacteria that cause infection of the urinary tract and sexual transmitted infections Upon completion of this course, the student will be able to: 1. List examples of urinary and genital tract specimens. 2. Explain methods of urine sampling and genital specimen collection for microbiology examination 3. Explain the use of the primary plating media for each specimen. 4. Describe urine culture technique for the diagnosis of UTI 5. Select incubation atmosphere, temperature, and time for each culture. 6. Interpret, and evaluate direct Gram stains of genital specimens. 7. Describe colonial morphology and growth characteristics of the bacteria 8. Determine appropriate biochemical tests or adjunct procedures required for identification of significant isolates. 9. Interpretation antibiotic susceptibility tests as required and evaluate their appropriateness with regard to the treatment 10. Explain microscopic examination for diagnose Treponema pallidum infection 11. Explain the procedure and principle laboratory diagnosis of Chlamydia trachomatis infection : microscopic, direct antigen detection, PCR 12. Correlate culture results with clinical history and presentation.
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III. Methods Presentation Demonstration Discussion

Laboratory Activity The laboratory examination is an integral part of the Microbiology course. It provides the student an opportunity to learn basic microbiological techniques, and introduces him/her to a case based approach to microorganisms related to respiratory tract infection. Steps in microbiology examinations: Direct detect infectious agent Visualization Culture Nucleic acid (DNA or RNA) Antigen detection Indirect detect hosts response to infection Serological detect antibody

Direct detection: o Visualization (Microscopic) - smears from lesions o Light microscope and Gram stain, wet mount preparation Bacteria Fungal spores o Electron microscope Viruses o Quick and easy if positive , negative results not definitive Visualize bacteria: secretion : genital discharge mucosal smears : mucosal scraping Direct detection: Culture and identification Growth medium : colony count, selective media Adequate incubations conditions Temperature Atmosphere (O2, CO2) Biochemical tests for identification Expensive and time-consuming, positive results may be definitive Inoculate enrichment or selective media, incubate 24 hours Streak on agar plates, incubate 24 hrs Pick presumptive colonies and restreak for identification
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METHODS OF URINE COLLECTION 1. Random collection taken at any time of day with no precautions regarding contamination. The sample may be dilute, isotonic, or hypertonic and may contain white cells, bacteria, and squamous epithelium as contaminants. In females, the specimen may contain vaginal contaminants such as trichomonas, yeast, and during menses, red cells. 2. Early morning collection of the sample before ingestion of any fluid. This is usually hypertonic and reflects the ability of the kidney to concentrate urine during dehydration which occurs overnight. If all fluid ingestion has been avoided since 6 p.m. the previous day, the specific gravity usually exceeds 1.022 in healthy individuals. 3. Clean-catch, midstream urine specimen collected after cleansing the external urethral meatus. 4. Catherization of the bladder through the urethra for urine collection is carried out only in special circumstances, i.e., in a comatose or confused patient. This procedure risks introducing infection and traumatizing the urethra and bladder, thus producing iatrogenic infection or hematuria. 5. Suprapubic transabdominal needle aspiration of the bladder. When done under ideal conditions, this provides the purest sampling of bladder urine. This is a good method for infants and small children. Method of clean-catch, midstream urine specimen collection: 1. Local disinfection of the meatus and adjacent mucosa should be performed with a nonfoaming antiseptic solution; this region should then be dried with a sterile swab to avoid mixture of the antiseptic with urine. 2. Contact of the urinary stream with the mucosa should be minimized by spreading the labia in females and by pulling back the foreskin in uncircumcised males. 3. A midstream urine is one in which the first half of the bladder urine is discarded and the collection vessel is introduced into the urinary stream to catch the last half. The first voided specimen should be discarded since the initial urine flushes urethral contaminants such as contaminating cells and microbes from the outer urethra. It is the second, midstream sample that should be sent to the laboratory. This sounds easy, but it isn't (try it yourself before criticizing the patient). Urine collection of baby

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URINE CULTURE 1. QUALITATIVE URINE CULTURE As previously discussed, the most common cause of UTI in men and women are gram negative rods from the patients fecal flora; therefore, a qualitative approa ch in demonstrating their presence should center around growth characteristics and biochemistry associated with that group. The Qualitative isolation of urinary pathogens begins with primary culture on Blood agar. Most all incriminating microorganisms will grow voraciously on sheep blood with E. coli representing 80% of UTI, and many producing beta hemolytic zones. Mac Conkey agar is utilized to further selectively isolate and differentiate those gram negative rods that ferment lactose (e.g. E.coli, Klebsiella, Enterobacter and Serratia). Patients who have high numbers of cells/ ml that grows on blood agar but not on MacConkey s agar may suggest a more serious invasive pathogen of the descending route (e.g. S. pyogenes , Candida or S. aureus). CULTURE PROCEDURE Step 1. Label the bottom of a blood agar and MacConkey agar plate Qualitative Urine Step 2. From the simulated urine, streak for isolation onto the blood and MacConkeys agar.
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Step 3. Incubate plates inverted at 370C for 24 hours. Step 4. Record observations in results

2. QUANTITATIVE URINE CULTURE a. Pour plate method : isolation of bacteria by dilution techniques b. Calibratied loop inoculation A quantitative culture can be easily performed using a 1:1000 ml (0.001 ml) sterile, disposable loop. The loop is dipped vertically into the mixed urine specimen and inoculated onto a blood agar plate making a single line streak from the top of the plate to the bottom of the plate. Distribution of the 1:1000 ml urine dilution is done by cross-streaking the initial line of inoculation over the surface of the plate to obtain countable colonies. The plate is incubated overnight at 35 0 C and examined the next day for countable colonies. The number of colonies present is multiplied by 1000 (1000 represents the reciprocal of your dilution factor RDF i.e. from the standardized loop 1/1000). Multiplying the number counted NC by the (RDF) gives the number of organisms/ml of urine. CFU/ml = NC x RDF CULTURE PROCEDURE Step. 1. Label the bottom of a blood agar plate Quantitative Urine Step 2. Each desk is provided with 1/1000 ml standardized inoculating loops. Aseptically remove one loop from the container Step 3. Using the standardized loop, recover 1/1000 ml of urine from the simulate specimen provided. Step 4. Perform a quantitative culture streak Step 5. Incubate plate at 370C for 24 hr. Step 6. Record CFU/ml

c. Dip slide method The functional unit consists of a clear plastic dual-chambered "paddle" containing both a general purpose and a differential medium, Trypticase soy agar (TSA) and Levine eosin methylene blue (EMB) agar or other kind of media, respectively. It is housed in a clear plastic screw-cap vial. The media beds are parallel and face the same direction; each has a surface size of about 310 mm2. The outer wall of the media wells is constructed to prevent medium dislodgement during handling and shipment and the trapping of urine between the wall and media. To inoculate the unit, the media portion is momentarily dipped into the urine to wet the entire agar surfaces. It is then returned to the vial and incubated in an upright position overnight at 370C. The number of colonies per milliliter of urine is estimated by comparing the density of growth with a standard chart obtained.
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Microorganisms causing urinary tract infections The bacteria that cause UTI will vary according to the clinical and epidemiological setting are usually originating from the patients own normal intestinal organisms; colonic flora (coliforms = Enterobacteriaceae) which, in certain circumstances, are able to pass from the area around the anus into the urethra and so into the bladder. Members of this bacterial group include opportunistic pathogens (e.g. E.coli by far the most frequent cause of urinary tract infection), Klebsiella, Enterobacter, Serratia, and Proteus. Pseudomonas and Enterococcus are also often incriminated, especially in hospitalized patients with indwelling urinary catheters. Community acquired UTI, otherwise healthy adults Escherichia coli most common Staphylococcus saprophyticus (usually young adult females) Less commonly: Proteus species Enterobacter species Serratia species Morganella species Klebsiella species Citrobacter species Pseudomonas aeruginosa Enterococcus species

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(Patients with recurrent UTIs, urinary tract abnormalities, obstruction and neurogenic bladder will have a higher frequency of those bacteria listed as less common above). Hospital acquired UTI Hospitalized patients rapidly become colonized by the hospitals resident microflora, specifically Gram negative bacteria. Patients in this setting are prone to UTI because they are debilitated and may have had urinary tract instrumentation, particularly urinary tract catheterization. Indwelling urinary catheters universally become colonized by bacteria. Hospital bacteria have been selected by broad spectrum antibiotics used in the hospital, and are generally quite resistant to antibiotics. Causes of hospital acquired UTIs include: E. coli and Enterobacteriaceae as described above. Increased frequency of Proteus, Enterobacter, Serratia, Morganella. Pseudomonas aeruginosa and other Pseudomonas species. Enterococcus faecalis and E. faecium. Stenotrophomonas maltophilia Corynebacterium urealyticum Staphylococcus epidermidis Candida albicans and other species. Uncommon causes of UTI Some other microorganisms which may infect the urinary tract should be mentioned: Mycobacterium tuberculosis renal TB; uncommon in this community. Adenovirus type 11 haemorrhagic cystitis in children. SEXUALLY TRANSMITTED INFECTIONS N. gonorrhoeae Specimen collection : Dacron or rayon swab Specimen transport 1. Direct inoculation ~ Candle jars 2. Nonnutritive transport media ~Amies medium (12 hrs) should not be refrigerated 3. Nutritive transport systems ~ Transgrow, JEMBEC, Bio-Bag, Gono-Pak Transgrow Medium is a convenient flask containing MTM Chocolate agar and CO 2. Culture Selective Media for N. gonorrhoeae are Thayer-Martin (Modified=MTM), New York City Medium (NYCM) Incubation condition 35oC, 3-5% CO2 Colonial morphology : 5 types autolysin
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N. gonorrhoeae forms small, convex, smooth, grayish-white to colorless, mucoid colonies in 48 hours at 35-370C. The gonococcus requires: enriched medium with increased CO2 tension for growth Cultured on modified Thayer Martin (MTM) Chocolate agar.

MTM Chocolate agar is selective for pathogenic Neisseria contains : enrichment factors to promote the growth of gonococci, antibiotics to inhibit normal body flora : vancomycin to inhibit gram-positive bacteria; colistin to inhibit gram-negative bacteria; trimethoprim to suppress Proteus; nystatin to inhibit yeast. Identification tests 1. Oxidase test tetramethyl--phenylenediamine dihydrochloride Oxidase indophenol (blue)

2. Carbohydrate utilization tests Cystine trypticase agar (CTA) ~ 1% carbohydrate, phenol red Organism N. gonorrhoeae N. meningitides N. lactamica Chlamydia trachomatis Chlamydia testing is available by 3 methods: Culture : cell culture Antigen detection from genital specimen: DFA. Elisa Molecular technique: Molecular Probe, PCR Chlamydia Culture Collection 1. Utilizing a sterile swab (rayon or dacron), obtain a suitable specimen. 2. Place the swab in transport media (M4 media tube with pink liquid). 3. Label the transport media vial. 4. Transport the vial to the laboratory. May be refrigerated. Chlamydia DFA/ Elisa Collection Glucose + + + Maltose + + Lactose + Sucrose -

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The Direct Fluorescent Antibody (DFA) method for Chlamydia utilizes an antibody to detect Chlamydia organisms on a slide, while Elisa detect chlamydia antigen using microtiter plate. This method can be used for urethral, cervical, and rectal samples. Urethral samples (male): Patient should not have urinated one hour prior to sampling. 1. Insert small dacron swab (mini-tip) 2-4 cm into urethra. 2. Rotate swab and withdraw. Cervical samples (swab): 1. Wipe exocervix with large or small rayon or dacron swab to remove all excess mucus. Dispose of swab. 2. Gently insert rayon or dacron swabs into endocervical canal until most of dacron tip is not visible. 3. Rotate swab 5-10 seconds inside endocervical canal. 4. Withdraw swab without touching any vaginal surfaces.

Rectal samples: Samples should be collected only from symptomatic patients. 1. Insert the large dacron swab about 3 cm into anal canal. 2. Move swab from side to side to sample crypts. 3. Withdraw swab. If fecal contamination occurs, discard swab and obtain another specimen. Treponema pallidum Laboratory Diagnosis: This relies heavily on clinical manifestations. In addition, the finding of Treponema pallidum within exudative lesions by the use of dark-field microscopy and positive serology aids the diagnosis. Penicillin treatment eradicates all stages, including congenital infection in pregnancy. PROCEDURE Step 1. Observe Prepared Slide Treponema pallidum. Special stain or negative stain (bright field microscope) Step 2. Observe Dark Field Microscopy (presumptive Identification) Step 3. Record results TERMS AND QUESTIONS FOR STUDY UTI exercise 1. What is bacteriuria? 2. How do microorganisms enter the urinary tract? 3. What group of microorganisms appear in ascending UTIs. Neisseria gonorrhea exercise
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1. 2. 3. 4.

Gram stain Bacteria isolation, media Colony morphology Identification procedure

Spirochaeta exercise 1. Special stain 2. Differentiate morphology of: Treponema pallidum Borrelia bergdorferi Leptospira interrogans ADDITIONAL QUESTION 1. What is the significance of small pin point, beta hemolytic, and colonies appearing on blood agar but no growth on MacConkey agar when conducting a qualitative urine culture? 2. What are significant numbers for a UTI when conducting a quantitative urine culture? How many CFU/ml would you conclude the number of colonies if you find 145 colonies on the plate when you inoculate using a 1/1000 ml calibrated loop? What number of colonies if you use a 1/100 ml calibrated loop? 3. Why is urine specimens keep cold until plated? 4. What do we learn from a dip stick test? Is this test conclusive? 5. Can we distinguish between N. gonorrhoeae and N. meningitidis by Gram stain? 6. What is the meaning of intracellular Gram negative cocci? Explain! 7. Why are selective media needed in primary isolation from female urogenital specimens? 8. Can we diagnose syphilis from Gram stained smears of primary lesions? Explain! 9. Explain dark field microscopy and its application in the Microbiology laboratory! DRAW THE RESULT OF YOUR ACTIVITY

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Signature of trainer -----------ss-------------

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WEEK VIII

HISTOLOGY AND PATHOLOGY ANATOMY OF URETER, URINARY BLADDER, URETHRA, AND EXTERNAL GENITALIA
LEARNING OBJECTIVES After performing laboratory activity, the student should be able to: 1. Describe the structure and function of ureters, urinary bladder, and urethra. 2. Compare the urethras of males and females. 3. Describe the pathogenesis and pathology of cystitis. 4. Describe the pathogenesis and pathology of urothelial carcinoma of the bladder. 5. Describe the pathogenesis and pathology of benign prostatic hyperplasia. 6. Describe the pathogenesis and pathology of prostatic adenocarcinoma. 7. Compare the prostate, seminal vesicles, and bulbourethral glands in terms of general organization, epithelial lining, secretory products, and the point(s) at which their secretions enter the excretory pathway. 8. Describe the histology of penis and female external genitalia 9. Describe the pathogenesis and pathology of cryptorchidism 10. Describe the pathogenesis and pathology of seminoma of the testes 11. Describe the pathogenesis and pathology of condyloma acuminate in penile 12. Describe the pathogenesis and pathology of squamous cell carcinoma

RESOURCE PERSON 1. Nursiah Nasution, dr 2. Hasrayati Agustina, dr., SpPA 3. Hermin Aminah, dr.,SpPA

REFERENCES 1. Sadler, TW. Langmans Medical Embryology. 10thEd. Lippincott Williams & Wilkins. Philadelphia, Baltimore, New York. 2006. pp.229-256. 2. Moore KL and Dalley AF. Clinically Oriented Anatomy. 5th Edition. Lippincott Williams & Wilkins. 2006 pp 405 - 414.
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3. Janqueira, L., Carlos and Carneiro. Basic Histology. Text and Atlas. 10th Ed. Angr Medical Books McGraw-Hill. pp 383-401, 431- 447. 4. Kumar V, Abbas AK and Fausto N. Robbins and Cotran Pathologic Basis of Diseases (2005), 7th Edition. Elsevier Saunders. pp 1024-1056.

INTRODUCTION The genital organs of man include the testes, epididymides, ductus deferentes, ejaculatory ducts, prostate, seminal glands and bulbourethral glands, also several supporting structures, including the scrotum and the penis. The female genital organs include the ovaries, the uterine tubes, the uterus, the vagina, and the external organ (vulva and pudendum). The female external genitalia, or vulva, consist of the clitoris, labia minora, labia majora, and some glands that open into the vestibulum, a space enclosed by the labia minora. Penis and scrotum are the male external genitalia. During laboratory activity, the students will be asked to show any anatomical parts using model, interactive CD, as well as histological preparations and finally also discuss about important aspects of physiology and pathogenesis. Before activity, the students have to accomplish the homework assignment first, read the primary references, so that they will be more readily performing laboratory activity

HOMEWORK ASSIGNMENT 1. HISTOLOGY

Key words:

Central zone Ejaculatory ducts Penile urethra Peripheral zone Prostatic urethra Seminal vesicle Transitional zone - Vas deferens -

Fig. 3.1 Organization of the prostate gland.

Fig. Prostate gland with prostatic urethra. Stain: H&E. 80x Key3.2 words : 90 Dilatation of utriculus Diverticulum of urethral wall Ejaculatory ducts Fibromuscular stroma - Prostatic glands (alveoli) - Prostatic urethra - Smooth muscle of the stroma - Utriculus

Prostate Prostate Gland Gland

1. Prostatic concretions

4. Glandular epithelium

Key words:
- Capillary and venules - Corpus amylaceum (prostatic concretions) - Glandular alveoli - Glandular ducts - Glandular epithelium - Smooth muscle fibers

5. Ducts 2. Glandular alveoli 6. Smooth muscle fibers

3. Glandular ducts 7. Capillary and venule

Fig.3.3 Prostate gland (sectional view). Stain: H&E. 180x

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Key words:
Adipose tissue Adventitia Artery Circular muscle layer Lamina propria Longitudinal muscle layer Lumen Nerves Transitional epithelium (basal layer) Transitional epithelium (superficial layer) Venules

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Fig. 3.4 Ureter (tansverse section). Stain: H&E. 50x.

Key words: - Adventitia - Transitional epithelium - Circular muscle layer - Longitudinal muscle layer - Basal layer of epithelial cells - Lamina propria

Fig. 3.5 Ureter (tansverse section). A section of the wall. Stain: H&E. 50x.

Fig. 3.6 Urinery bladder, upper part: (tansverse section). A section of the wall. Stain: H&E. 40x.
Key words: Interstitial connective tissue Lamina propria Serosa Smooth muscle bundles Transitional epithelium lining the bladder

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KEY ANSWER: - Cavities (cavernous veins) of corpus cavernosum

- Central (deep) artery - Corpus cavernosum - Corpus spongiosum Think first tunic and answer the question correctly and concisely! - Dartos -1. Deep penile fascia Explain the epithelum lining part of the penile urethra. -2. Dermis - Dorsal artery 4. PATHOLOGY ANATOMY - Epidermis

Medial septum Sebaceous gland in dermis Superficial vein Trabeculae Tunica albigunea of corpus cavernosum Tunica albigunea of corpus spongiosum Urethra

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Testis

Figure: testis

Please complete this figure by naming the pointed parts!

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SEMINIFEROUS TUBULES, STRAIGHT TUBULES, RETE TESTIS AND DUCTULI EFFERENTES

1. Seminiferous tubules

3. Connective tissue of mediastinum 4. Rete testis tubules 5. Ductuli efferentes (efferent ductules 6. Rete testis tubules

2. Straight tubules

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Please complete this figure by naming the pointed parts!

Ductus Ductus Epididymidis Epididymidis (Duct (Duct of of the the Epididymis) Epididymis)
1. Connective tissue 2. Cross sections of the ductus epididymidis 5. Section through of Ubend of the ductus epididymidis

6. Epididymal wall cut tangentially 3. Basement membrane 7. Smooth muscle fibers 8. Stereocilia 4. Pseudostratified columnar epithelium with stereocilia 9. Columnar cells 10. Basal cell

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Seminal Seminal Vesicle Vesicle

4. Glandular epithelium 1. Crypts in the mucosa

5. Primary fold in the mucosa 6. Secondary folds 7. Lamina propria

2. Muscular coat 3. Adventitia

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Please complete this figure by naming the pointed parts!

Ductus Ductus Deferens Deferens (Transverse (Transverse section) section)

1. Outer longitudinal muscle layer

2. Circular muscle layer 3. Inner longitudinal muscle layer

5. Lamina propria 6. Longitudinal crest of lamina propria 7. Epithelium

8. Adipose tissue

4. Nerve and blood vessels in the adventitious

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Think first and answer the question correctly and concisely! 1. 2. 3. Describe the location of the gland, ducts, and external genitalia of the male reproductive system Describe the general organization of the testis Describe the differences in wall structure of excretory genital ducts.

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3. PATHOLOGY ANATOMY

1. CHRONIC NON SPECIFIC CYSTITIS AND CYSTITIS CYSTICA

A 37 years old female came to the outpatient clinic because of disuria and frequency since 2 weeks ago .She had no fever or flank pain

Macroscopic

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Microscopic:

2. UROTHELIAL CARCINOMA OF THE BLADDER

A 56-year-old male had several episodes of hematuria and dysuria in the past few week. The patient was a chronic smoker for 30 years. A urinalysis show 4 (+) blood. The urine culture is negative. A cystoscopy was performed, and a 2 cm, sessile, friable mass was seen on the left lateral wall.

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Macroscopic:

Microscopic :

3. BENIGN PROSTATIC HYPERPLASIA

A 65-year-old male had increasing difficulty in urination. He had to get up several times at night because of a feeling of urgency. He has difficulty starting and stopping urination. His serum PSA level was slighty increased.
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Macroscopic:

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Microscopic:

4. PROSTATIC ADENOCARCINOMA
A 65-year-old male presented with a long history of urinary hesitancy nocturia. Digital rectal examination revealed a hard, irregular prostate. Serum PSA level is 120 mg/ml.

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Macroscopic:

Microscopic:

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Answer these questions correctly!! 5. CRYPTORCHIDISM A 2 year-old boy was brought to the pediatric outpatient clinic because his parent noticed that he has testes in his right scrotum

Microscopic :

6. SEMINOMA OF THE TESTES

A 57 year-old male had a unilateral enlargement of the testss. Physical exanination showed enlargement of the left testes 5 cm in diameter

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Macroscopic :

Microscopic :

7. CONDYLOMA ACUMINATA PENIS

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Microscopic :

8. SQUAMOUS CELL CARCINOMA PENIS

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Microscopic :

HOMEWORK ASSIGNMENT A. Fill in the blank box under each the picture with microscopic and macroscopic appearance of each cases. B. Answer these questions correctly!! 1. Describe the pathogenesis of cystitis. 2. Please explain about congenital malformation of urinary bladder. 3. Describe the pathogenesis of urothelial carcinoma of the bladder. 4. Please explain about urethral inflammation. 5. Describe the pathogenesis of benign prostatic hyperplasia. 6. Describe the pathogenesis of prostatic adenocarcinoma. 7. Please explain about congenital anomaly of penis 8. Describe the pathogenesis of cryptorchidism 9. Please explain about testicular inflammation 10. Describe the pathogenesis of seminoma of the testes 11. Describe the pathogenesis and pathology of condyloma acuminate in penile 12. Describe the pathogenesis and pathology of squamous cell carcinoma

LABORATORY ACTIVITY Pre-requisites: The students have to do the homework assignment and read the references as listed in the page above. The class will be divided into 4 groups performing 4 main activities
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1. Histology a. Discus the homework materials b. See the specimen under microscope and try to make a schematic draw and put the most important description based on the schematic draw. Specimen No.1 Schematic Draw of Description

Specimen No.2

Schematic Draw of

Description

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Specimen No.3 Schematic Draw of Description

Specimen No.4 Schematic Draw Description

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Specimen No.5 Schematic Draw of Description

2. Pathology a. Discus the homework materials b. See the specimen under microscope and try to make a schematic draw and put the most important description based on the schematic draw.

Specimen No 1: Schematic Draw

Description

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Specimen No 2: Schematic Draw

Description

Specimen No 3: Schematic Draw

Description

Specimen No 1: Schematic Draw

Description

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Specimen No 2: Schematic Draw

Description

Specimen No 3: Schematic Draw

Description

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Specimen No 4: Schematic Draw

Description

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