Clinical Pathology and Bone Marrow Evaluation Study Guide

Overview of Clinical Pathology and Clinical Skills

• Clinical pathology consists of several key components: blood smears, urinalysis, and clinical chemistry.

• Clinical chemistry focuses on the analysis of electrolytes, proteins, and waste products.

• Clinical skills education involves hands-on laboratory work rather than just lectures. Performance is evaluated through clinical skills exams, which incorporate elements of clinical pathology and potentially toxicology.

• Upcoming laboratory activities on the 5th floor include:

  • Collecting blood samples.

  • Dividing into groups to create blood smears.

  • Identifying cellular components of blood: red cells, neutrophils, monocytes, and lymphocytes.

  • Learning how to count cells and understand their clinical significance.

  • Measuring Packed Cell Volume (PCV).

Case Study: 11-Year-Old English Springer Spaniel (Piper)

• Patient Profile: 11-year-old spayed female English Springer Spaniel named Piper.

• Clinical History:

  • Presented with lymphadenopathy (enlarged lymph nodes).

  • History of mild nephronomy.

  • Swelling above the right thigh.

  • Tested positive for Lyme disease six months ago and was treated with doxycycline.

  • Geographic association: Lyme disease is primarily associated with the Eastern part of the country (East Coast).

• Physical Examination Findings:

  • Petechiae and ecchymosis present on the tongue and mucous membranes.

  • Bounding pulses.

  • Enlarged lymph nodes.

Hemostasis and Mucocutaneous Lesions

• Petechiae: These are described as little red dots, similar to pinheads or pinpoint hemorrhages. In hematology, petechiae are an indicator of issues with primary hemostasis, specifically involving platelets.

• Ecchymosis: These are larger lesions, similar to a bruise or a paintbrush appearance. They indicate more serious processes than pinpoint hemorrhages.

• Kissing Lesion: This is a specific dental and mucous membrane finding in dogs. It appears as a lesion on the mucous membrane of the lip that mirrors an ulcer on the opposing part of the gum or tooth. Unlike ecchymosis, a kissing lesion is caused by the proximity and contact of the lip with a diseased tooth or ulcerated gum area.

Clinical Hematology: Data Interpretation and Reference Ranges

• Blood work results are generally divided into three cellular groups: myeloid (white blood cells), erythroid (red blood cells), and thrombocytes (platelets).

• Reference Ranges: These are specific to species and sometimes breeds. They vary by laboratory; a reference range from one lab should never be applied to results from another lab.

• Absolute Numbers vs. Percentages: The absolute number is the only valid metric for interpreting blood work (e.g., determining neutrophilia). Percentages are used only as a step to calculate absolute numbers. Relying on percentages for diagnosis is clinically incorrect.

• Quantifying Changes: In clinical pathology, increases or decreases are described using specific qualifiers:

  • Mild increase/decrease.

  • Moderate increase/decrease.

  • Marked increase/decrease.

• Relative changes are often expressed as "folds" (e.g., a twofold, threefold, or tenfold increase).

• Avoid using the term "elevation" when describing cell counts; use "increase" or "decrease" instead.

The Three Cellular Lineages of Blood

• Myeloid Component: Includes segmented neutrophils, band neutrophils, lymphocytes, monocytes, eosinophils, and basophils.

• Erythroid Component: Red blood cell mass calculated via red blood cell count, hemoglobin, and hematocrit/PCV.

• Thrombocyte Component: Platelets, which are essential for clotting.

• Cytopenias:

  • Bicytopenia: A reduction in the absolute numbers of two of the three cellular lines.

  • Pancytopenia: A reduction in the absolute numbers of all three cellular lines (myeloid, erythroid, and thrombocyte).

• Mechanisms of Cytopenia: When cell numbers decrease, four possibilities must be considered:

  1. The cells are being destroyed.

  2. The cells are being used up (consumption).

  3. The cells are being lost (hemorrhage).

  4. The cells are not being produced in the bone marrow (the "factory").

Hematocrit and Packed Cell Volume (PCV)

• Packed Cell Volume (PCV): This is a manual measurement of the percentage of red blood cells in a sample. It is performed by filling a capillary tube with blood, spinning it in a centrifuge, and measuring the fractions. PCV is considered the most accurate measure of red cell mass.

• Hematocrit: This is a calculated value provided by automated machines.

• Nucleated Red Cells: In species such as birds (parrots/macaws), reptiles (snakes), and amphibians, red blood cells are nucleated. In these species, automated hematocrit calculations are inaccurate; PCV must be measured manually using the capillary system.

Bone Marrow Evaluation: Aspirates and Biopsies

• Bone Marrow Aspirate:

  • Involves taking a fluid sample to create a smear for cytology.

  • Quick and relatively inexpensive (approximately $600-800$ dollars, including sedation).

  • Provides rapid results (often within 24 hours).

• Bone Marrow Biopsy:

  • Involves taking a tissue core for histopathology.

  • Provides a superior picture of the bone marrow "in situ," showing the actual structure and architecture.

  • More expensive (approximately $1000-1500$ dollars) and takes longer for results (roughly 72 hours).

• Sampling Sites:

  • Costochondral Junction: Applicable for most species, including foals.

  • Sternum: Used in adult animals.

  • Wing of Ilium: Common site for dogs and foals.

  • Humerus: Used in adults or obese patients; should be avoided in young animals.

  • Femur: Used in cats and small dogs.

• Procedural Risks: The primary risk is bleeding, especially in patients with thrombocytopenia. This must be discussed with clients and documented as part of a mitigation strategy.

Indications for Bone Marrow Investigation

• Persistent Abnormalities: Bone marrow should be investigated if peripheral blood abnormalities (such as anemia or leukopenia) are persistent. "Persistent" implies that the condition remains unchanged over several weeks (e.g., 3 to 6 weeks) despite treatment.

• Hematopoiesis Timeline: The process of forming cellular components takes time; it can take up to 40 days for the entire process, and around 14 days (2 weeks) to see significant changes in peripheral circulation after the bone marrow responds.

• Atypical Cells: The presence of abnormal or immature cells in circulation is a strong indication for bone marrow study.

• Staging Neoplasia: Used to check for metastasis or primary bone marrow involvement in conditions like mast cell neoplasia, lymphoma, leukemia, or metastatic neoplasia.

• Myelophthisis: A process where the bone marrow is "crowded out" by abnormal cells, such as tumor cells. This is metaphorically described as a "fat man sitting on the precursor cells," leading to the death of normal cells and causing pancytopenia. Osteosarcoma is a common tumor that can metastasize and cause this.

• Detection of Microorganisms: Certain pathogens, such as Leishmania, can be identified in bone marrow aspirates.

Cytology: Cellular Description and Identification

• Nuclear to Cytoplasmic (N/C) Ratio: This describes the size of the nucleus relative to the amount of cytoplasm in a cell.

  • A high N/C ratio means the nucleus occupies most of the cell (common in blast cells or neoplastic cells).

  • A low N/C ratio means there is a significant amount of cytoplasm compared to the nucleus.

• Nuclear Placement:

  • Centrally placed: Nucleus is in the middle of the cell.

  • Eccentric: Nucleus is pushed to one side (e.g., in plasma cells).

• Cytoplasm Appearance: Described by color (deep bluish, light purple, pink) and texture.

• Round Cells: A general category of cells typically characterized by round shapes and often high N/C ratios.

Myeloid and Erythroid Lineages in Cytology

• Myeloid Series Identification: The nuclei in the myeloid lineage eventually become "squished" or "choked," taking on shapes like a horseshoe or a bow tie as they mature. The cytoplasm in these cells is often lighter or different in color compared to the erythroid series.

• Erythroid Series Identification: The nuclei remain round and resist the "choking" or "squishing" seen in myeloid cells. As erythroid cells mature, their nuclei become smaller, darker, and more condensed (pyknotic).

• Myeloid:Erythroid (M:E) Ratio: A calculation based on counting a specific number of cells (e.g., 10 or 100) to determine the balance of production in the bone marrow.

• Hyperplasia:

  • Erythroid Hyperplasia: An overproduction of red cell precursors, which can elbow out the myeloid lineage.

  • Myeloid Hyperplasia: An overproduction of white cell precursors.

Iron Storage and Species Differences

• Prussian Blue Stain: A specific stain used to identify iron (Fe) in bone marrow samples. Iron appears as dark spots or "iron stores."

• Absence of Iron: If no iron is seen in the bone marrow of certain species, it indicates iron deficiency anemia.

• Species Variation: While most species store visible iron in the bone marrow, cats are an exception. It is normal to see no visible iron stores in a feline bone marrow aspirate; their iron is primarily stored within macrophages.

Questions & Discussion

• Question: Are we going to be tested on the lab work?

• Answer: Students will be tested on what they perform during the clinical skills hours. This testing relates to clinical pathology and may include toxicology if it falls within the same timeframe. The goal is the integration of knowledge, not just memorization.

• Question: Regarding the risk of bleeding in thrombocytopenic patients, would we still perform a biopsy if we saw a mass on an organ like the kidney or liver?

• Answer: One must be extremely cautious. Biopsying a mass in a dog with low platelets risks the animal bleeding to death. The risk must be discussed with the client, a mitigation strategy plan must be created, and the discussion must be documented and signed.

• Question: What is the ratio of the nucleus to the cytoplasm in the cell on the right?

• Answer: It is about $1:1$. The cell on the right has a higher ratio compared to others. This cell can be described as a round cell with deep bluish cytoplasm and a high nuclear cytoplasmic ratio.

• Question: Is there a specific name for the process of cells dividing on the slide?

• Answer: These are called mitotic figures. Seeing many mitotic figures indicates actively dividing cells, which is normal in the bone marrow but can also be a hallmark of tumor cells.