Unit 4: Clinical Pathology & Immune System| Pathology | 4th Semester of Bachelor of Optometry

CLINICAL PATHOLOGY

Interpretation of Urine Report

Introduction:
Urinalysis is one of the oldest, simplest and most informative laboratory tests used in clinical practice. A routine urine report consists of three broad components — physical (macroscopic), chemical (dipstick), and microscopic examination — and provides crucial information about renal function, metabolic disorders (e.g., diabetes, ketonuria), urinary tract infections, and systemic diseases. For optometry students, urine reports are clinically relevant because systemic conditions detected on urinalysis (notably diabetes mellitus, hypertension-related renal disease, and infections) have direct implications for ocular health and patient management (e.g., diabetic retinopathy screening frequency, perioperative risk assessment).

Specimen Collection & Pre-analytical Considerations

Type: Random (spot) urine is most common; early-morning sample is concentrated and preferred for many tests (e.g., protein, microscopy).
Midstream clean-catch: Reduces contamination for culture and microscopy.
Catheter or suprapubic aspirate: Used when contamination must be avoided (infants, incontinent patients).
Timing: 24-hour urine collection is used for quantitative measurement (protein, creatinine clearance).
Handling: Analyze within 1 hour if possible; refrigerate at 4°C if delayed to avoid bacterial overgrowth or cell degeneration.

Stepwise Interpretation Approach (Practical)

Check patient details, collection method, and time of sample.
Review physical appearance: color, turbidity, odor.
Review dipstick (chemical) results and specific gravity.
Correlate dipstick positives with microscopy (e.g., dipstick blood vs RBCs on microscopy).
If infection suspected, correlate leukocyte esterase and nitrite with WBCs and bacteriuria; consider urine culture.
Consider clinical context (symptoms, known diseases like diabetes, pregnancy, medications).

Normal Urine Values (Typical Reference)

Color: pale yellow to amber
Clarity: clear
Specific gravity (SG): 1.005–1.030 (reflects concentrating ability)
pH: 4.5–8.0 (usually slightly acidic)
Protein (dipstick): negative or trace <30 mg/dL
Glucose: negative
Ketones: negative
Blood (dipstick): negative
Leukocyte esterase: negative
Nitrite: negative
Bilirubin and urobilinogen: negative or within low range
Microscopy: 0–2 RBCs/HPF, 0–5 WBCs/HPF, few squamous epithelial cells, no casts or crystals of clinical significance

Physical (Macroscopic) Examination

Color:
- Normal: pale yellow — amber (urobilin)
- Dark amber: concentrated urine, bilirubin (cholestasis), some drugs
- Red/pink: RBCs (hematuria), myoglobinuria, beetroot, porphyria
- Brown/tea-colored: hemoglobinuria, myoglobin, methemoglobinemia, some drugs
- Cloudy: pyuria, lipiduria, phosphaturia
Odour: Fruity aroma — ketonuria (diabetes); foul smell — infection.
Clarity/Turbidity: Clear normally; turbidity suggests cells, crystals, mucus, bacteria, or lipids.
Specific Gravity (SG): Indicates concentration. Low SG (close to plasma) suggests impaired concentrating ability (e.g., diabetes insipidus, renal failure). High SG suggests dehydration or presence of glucose/protein.

Chemical (Dipstick) Parameters — Meaning & Interpretation

pH

Normal 4.5–8.0. Acidic urine: high protein diet, dehydration. Alkaline urine: UTI with urease-producing organisms, vegetarian diet, prolonged storage (bacterial overgrowth).

Specific Gravity

Low (≈1.005): dilute urine — excessive fluid intake, diabetes insipidus. High (≈1.030): concentrated urine, glycosuria, SIADH.

Protein (Albumin)

Dipstick primarily detects albumin (not globulins).
Trace to 1+ may occur after exercise or fever (functional proteinuria). Persistent ≥1+ suggests renal disease (glomerulonephritis, diabetic nephropathy, nephrotic syndrome if massive proteinuria >3.5 g/24 hr).
Confirm with urine albumin-to-creatinine ratio (ACR) or 24-hr protein.

Blood (Hematuria) — Dipstick

Detects heme—positive with RBCs, hemoglobin, or myoglobin. Correlate with microscopy:
RBCs on microscopy → true hematuria (stones, infection, malignancy, glomerular disease).
No RBCs but positive dipstick → hemoglobinuria (intravascular hemolysis) or myoglobinuria (rhabdomyolysis).

Glucose

Normally negative. Positive glucose indicates glycosuria — commonly due to hyperglycemia (diabetes mellitus) when plasma glucose exceeds renal threshold (~180 mg/dL). Also seen with proximal renal tubular defects (renal glycosuria).

Ketones

Positive in uncontrolled diabetes (DKA), starvation, prolonged vomiting, or following ketogenic diets. Presence warrants urgent assessment in diabetic patients.

Bilirubin & Urobilinogen

Bilirubin positive suggests conjugated hyperbilirubinemia (obstructive or hepatocellular jaundice). Urobilinogen increased in hemolysis or hepatic dysfunction.

Nitrite

Many Gram-negative bacteria (e.g., E. coli) convert nitrate to nitrite — a positive nitrite suggests bacterial UTI (sensitivity depends on organism & urine dwell time).

Leukocyte Esterase

Produced by neutrophils; a positive test suggests pyuria and possible UTI. Should be interpreted with nitrite and microscopy.

Microscopic Examination — Elements & Clinical Correlation

Cells

Red blood cells (RBCs): 0–2/HPF normal. Dysmorphic RBCs and RBC casts suggest glomerular origin (glomerulonephritis). Isomorphic RBCs suggest lower urinary tract source (stones, tumor, infection).
White blood cells (WBCs): 0–5/HPF normal. Increased WBCs (>10/HPF) suggest infection or inflammation (pyelonephritis, cystitis). WBC casts indicate renal parenchymal infection.
Squamous epithelial cells: Common contaminant from distal urethra or vagina if numerous; few are normal.
Renal tubular epithelial cells: Suggest tubular injury (acute tubular necrosis, toxin damage).

Casts

Casts are formed in renal tubules and indicate renal origin:

Hyaline casts: May be normal after exercise or dehydration.
RBC casts: Pathognomonic of glomerulonephritis.
WBC casts: Suggestive of pyelonephritis or interstitial nephritis.
Granular casts: Nonspecific — seen in ATN or chronic kidney disease.
Fatty casts / oval fat bodies: Nephrotic syndrome (lipiduria).
Broad waxy casts: Chronic renal failure / advanced CKD.

Crystals

Common crystals: calcium oxalate (envelope-shaped), uric acid (rhombic), triple phosphate / struvite (coffin-lid, associated with alkaline urine and infection), cystine (hexagonal — cystinuria).
Significance: some crystals are common and benign; others indicate metabolic disease or predisposition to stones (e.g., uric acid, cystine).

Microorganisms & Others

Bacteria: presence with WBCs and positive nitrite/leukocyte esterase suggests UTI; confirm with urine culture for species and sensitivity.
Yeast: Candida — common in diabetics or catheterized patients.
Parasites: Schistosoma haematobium (terminal hematuria in endemic areas).

Common Urine Report Patterns & Their Interpretation

Glycosuria + Ketones + Hyperglycemia: Suggests uncontrolled diabetes mellitus — risk of diabetic ketoacidosis (DKA) if ketones high.
Protein (≥1+) + RBC casts + Dysmorphic RBCs: Strongly suggest glomerulonephritis — urgent nephrology referral.
Proteinuria (3+), oval fat bodies, hypoalbuminemia: Suggest nephrotic syndrome (massive protein loss).
Leukocyte esterase + nitrite + pyuria + bacteriuria: Urinary tract infection — obtain culture if symptomatic or complicated.
Isolated microscopic hematuria with normal dipstick: Investigate for stones, malignancy, or glomerular disease depending on age and risk factors.
High specific gravity with normal glucose/protein: Dehydration or concentrated urine — advise hydration and repeat.

Special Tests & Quantitative Measures

Urine albumin-to-creatinine ratio (ACR): Preferred screening test for diabetic nephropathy. Microalbuminuria: ACR 30–300 mg/g; macroalbuminuria >300 mg/g.
24-hour urine protein: Quantifies proteinuria (nephrotic range >3.5 g/day).
Urine culture & sensitivity: Gold standard for diagnosing UTI and guiding antibiotics.
Urine cytology: For suspected urothelial carcinoma with persistent hematuria.

Clinical Relevance to Optometry

Diabetes mellitus: Glycosuria and microalbuminuria indicate poor glycemic control and early diabetic nephropathy; such patients require more frequent diabetic retinopathy screening and perioperative assessment before ocular surgery.
Hypertension & renal disease: Proteinuria and reduced concentrating ability suggest renal involvement from hypertension — systemic control affects ocular perfusion and glaucoma risk.
Systemic infection / sepsis: Positive nitrite/leukocyte esterase with systemic signs may affect ophthalmic surgical planning and infection control.
Drug toxicity / rhabdomyolysis: Myoglobinuria (dipstick positive for blood without RBCs) can follow certain drugs or muscle breakdown; systemic management is primary.
Preoperative evaluation: Abnormal urine tests in diabetic or hypertensive patients may prompt delay or modification of ocular procedures and multidisciplinary management.

Interpretation of Blood Smears

Introduction:
Peripheral blood smear (PBS) examination is one of the most valuable investigations in hematology. Despite advanced automated analyzers, a well-prepared and properly interpreted blood smear remains indispensable for diagnosing anemia, infections, leukemias, parasitic diseases, and many systemic disorders. For optometry students, understanding blood smear interpretation is important because systemic hematological diseases often present with ocular manifestations such as conjunctival pallor, retinal hemorrhages, cotton wool spots, and papilledema. Thus, knowledge of blood smear findings aids in correlating systemic pathology with ocular findings.

Preparation of Blood Smear

Obtain capillary or venous blood.
Place a small drop near one end of a clean glass slide.
Use another slide (spreader) at 30–45° angle to spread the drop forward smoothly.
Smear should be tongue-shaped, thin at the feathered edge, and cover 2/3 of the slide.
Allow to air-dry and stain (commonly Leishman, Wright, or Giemsa stain).

Systematic Approach to Interpretation

Examine overall smear quality: Thickness, staining quality, distribution of cells.
Red Blood Cells (RBCs): Evaluate size, shape, color (hemoglobin content), inclusions.
White Blood Cells (WBCs): Count differential, assess morphology.
Platelets: Estimate number and size.
Look for parasites, abnormal cells, or other inclusions.

Red Blood Cell (RBC) Morphology

Normal: Normocytic, normochromic biconcave discs with central pallor.
Size abnormalities:
- Microcytes: small cells (iron deficiency anemia, thalassemia).
- Macrocytes: large cells (megaloblastic anemia, liver disease).
Color (Hemoglobin content):
- Hypochromia: increased central pallor (iron deficiency, thalassemia).
- Hyperchromia: rare, seen in spherocytosis.
Shape abnormalities (poikilocytosis):
- Spherocytes: hereditary spherocytosis, autoimmune hemolysis.
- Sickle cells: sickle cell anemia.
- Target cells: thalassemia, liver disease.
- Schistocytes (fragmented cells): DIC, TTP, microangiopathic hemolysis.
- Elliptocytes/ovalocytes: hereditary elliptocytosis, megaloblastic anemia.
- Tear drop cells: myelofibrosis.
Inclusions:
- Basophilic stippling: lead poisoning, thalassemia.
- Howell–Jolly bodies: post-splenectomy, megaloblastic anemia.
- Heinz bodies: G6PD deficiency.
- Malaria parasites: trophozoites, gametocytes inside RBCs.

White Blood Cell (WBC) Examination

Total and differential count: Relative proportion of neutrophils, lymphocytes, monocytes, eosinophils, basophils.
Neutrophils:
- Normal: multilobed nucleus, fine granules.
- Left shift: increased immature forms (band cells) in infections.
- Toxic granulation/Döhle bodies: severe infections, inflammation.
Lymphocytes:
- Reactive (atypical) lymphocytes: infectious mononucleosis.
- Small mature lymphocytes: seen in chronic lymphocytic leukemia.
Monocytes: Large cells with kidney-shaped nucleus; increase in chronic infections, leukemias.
Eosinophils: Bright orange granules; increased in allergies, parasitic infections.
Basophils: Deep purple granules; increase in CML, myeloproliferative disorders.
Malignant/abnormal cells:
- Blasts: large immature cells with high N:C ratio, fine chromatin, nucleoli (seen in acute leukemia).
- Auer rods: needle-like inclusions in AML blasts.

Platelet Evaluation

Normal platelets: 150,000–400,000/µL; appear as small purple cytoplasmic fragments.
Thrombocytopenia: reduced number — ITP, marrow failure, DIC.
Thrombocytosis: increased number — myeloproliferative disorders, reactive.
Giant platelets: in Bernard–Soulier syndrome, myeloproliferative states.

Parasitic & Infective Findings

Malaria: Ring forms, schizonts, or gametocytes inside RBCs.
Microfilaria: Motile organisms seen in peripheral blood at night.
Trypanosomes: Flagellated protozoa in plasma.
Leishmania (LD bodies): Intracellular in monocytes (rare in blood films, more in marrow).

Correlation with Clinical Disorders

Anemia

Iron deficiency: microcytic, hypochromic RBCs, target cells.
Megaloblastic: macrocytes, hypersegmented neutrophils.
Hemolytic: spherocytes, schistocytes, polychromasia.
Sickle cell anemia: sickled RBCs.

Leukemia

ALL: lymphoblasts.
AML: myeloblasts with Auer rods.
CLL: many mature small lymphocytes, smudge cells.
CML: leukocytosis with all stages of granulocytes (myelocytes, metamyelocytes, basophilia).

Infections

Acute bacterial: neutrophilia, left shift, toxic changes.
Viral: lymphocytosis, reactive lymphocytes.
Parasitic: eosinophilia, specific parasite visualization.

Ocular Relevance of Blood Smear Findings

Anemia: Conjunctival pallor, retinal hemorrhages, cotton wool spots, optic disc pallor.
Leukemia: Retinal hemorrhages, Roth’s spots, venous engorgement, optic nerve infiltration.
Thrombocytopenia: Subconjunctival hemorrhages, retinal hemorrhages.
Parasitic infections (e.g., malaria): Retinopathy with hemorrhages, cotton wool spots, papilledema.

Immune System

Introduction:
The immune system is the body’s defense mechanism against infections, tumors, and other harmful agents. It distinguishes “self” from “non-self” and orchestrates protective responses. A balanced immune system is vital for maintaining health, but dysfunction may lead to immunodeficiency, autoimmunity, hypersensitivity, or transplant rejection. For optometry students, understanding the immune system is crucial because many ocular diseases such as allergic conjunctivitis, uveitis, keratitis, dry eye disease, and autoimmune conditions like Sjögren’s syndrome and thyroid eye disease are immunologically mediated.

Components of the Immune System

1. Innate Immunity (Non-specific, First Line Defense)

Physical barriers: Skin, mucous membranes, conjunctiva, cornea, tear film.
Cells: Neutrophils, macrophages, dendritic cells, natural killer (NK) cells.
Molecules: Complement proteins, cytokines, lysozyme in tears.
Characteristics: Immediate, no memory, not antigen-specific.

2. Adaptive (Acquired) Immunity (Antigen-specific)

Humoral immunity: Mediated by B-lymphocytes producing antibodies (IgG, IgM, IgA, IgE, IgD).
Cell-mediated immunity: Mediated by T-lymphocytes (helper T-cells CD4+, cytotoxic T-cells CD8+, regulatory T-cells).
Characteristics: Antigen-specific, slower onset, has memory, improves on repeated exposure.

3. Lymphoid Organs

Primary lymphoid organs: Bone marrow (B-cell maturation), thymus (T-cell maturation).
Secondary lymphoid organs: Lymph nodes, spleen, mucosa-associated lymphoid tissue (MALT) including conjunctival-associated lymphoid tissue (CALT).

Cells of the Immune System

Lymphocytes: B-cells, T-cells, NK cells.
Antigen-presenting cells: Dendritic cells, macrophages, B-cells.
Phagocytes: Neutrophils, monocytes/macrophages — engulf and destroy microbes.
Accessory cells: Mast cells, basophils, eosinophils — mediate allergic and inflammatory responses.

Immune Mechanisms

Humoral Immunity

Mediated by antibodies secreted by plasma cells (derived from B-cells).
Neutralization of toxins/viruses, opsonization (enhancing phagocytosis), complement activation, and prevention of pathogen adherence.
Example: Protection after vaccination, allergic conjunctivitis (IgE-mediated).

Cell-mediated Immunity

Mediated by T-lymphocytes.
Helper T-cells (CD4+) regulate immune responses by cytokine secretion.
Cytotoxic T-cells (CD8+) kill virus-infected or tumor cells.
Example: Granulomatous uveitis in tuberculosis, rejection of corneal transplant.

Major Histocompatibility Complex (MHC)

MHC-I: Present on all nucleated cells; present peptides to CD8+ T-cells.
MHC-II: Present on antigen-presenting cells; present antigens to CD4+ T-cells.
Clinical importance: Tissue matching for transplantation, including corneal grafts.

Hypersensitivity Reactions (Gell & Coombs Classification)

Type I (Immediate, IgE-mediated): Allergic conjunctivitis, anaphylaxis.
Type II (Cytotoxic, antibody-mediated): Autoimmune hemolytic anemia, ocular cicatricial pemphigoid.
Type III (Immune complex-mediated): Vasculitis, systemic lupus erythematosus, peripheral ulcerative keratitis.
Type IV (Delayed, T-cell mediated): Contact dermatitis, phlyctenular keratoconjunctivitis, tuberculous uveitis.

Disorders of the Immune System

1. Immunodeficiency

Primary: Congenital (e.g., SCID, IgA deficiency).
Secondary: Acquired (e.g., HIV/AIDS, chemotherapy-induced).
Ocular relevance: Opportunistic infections (CMV retinitis, fungal keratitis).

2. Autoimmune Disorders

Immune response against self-antigens.
Examples: Rheumatoid arthritis, Sjögren’s syndrome, systemic lupus erythematosus, thyroid eye disease.
Ocular relevance: Keratoconjunctivitis sicca, scleritis, uveitis, optic neuritis.

3. Hypersensitivity/Allergic Disorders

Overactive immune response to harmless antigens.
Ocular examples: Seasonal/perennial allergic conjunctivitis, vernal keratoconjunctivitis, atopic keratoconjunctivitis.

4. Transplant Rejection

Immune-mediated damage to transplanted tissue.
Ocular relevance: Corneal graft rejection characterized by corneal edema, keratic precipitates, and Khodadoust line.

Laboratory Evaluation of Immune Function

Complete blood count with differential: Lymphopenia or eosinophilia suggests immune abnormalities.
Flow cytometry: CD4/CD8 counts in HIV.
Serology: Autoantibody tests (ANA, RF, anti-dsDNA, anti-SSA/SSB).
Immunoglobulin levels: IgG, IgA, IgM quantification.
Complement assays: C3, C4 levels in lupus.
Allergy testing: Skin prick test, serum IgE levels.

Ocular Immunology Highlights

Immune privilege of the eye: The eye limits inflammatory responses to preserve vision. Mechanisms include blood–ocular barriers, absence of lymphatics in cornea and anterior chamber, and local immunosuppressive microenvironment.
Anterior chamber–associated immune deviation (ACAID): Special immune tolerance induced when antigens are introduced into the anterior chamber.
Clinical importance: Reduced risk of rejection in corneal transplantation compared to other organs, though not absent.