L1 - INTRO TO HISTOLOGY, MICROSCOPY.pdf

ITM 101 | HISTOLOGY LESSON # 1 –INTRODUCTION TO HISTOLOGY 1st YEAR | A.Y. 2023-2024 | AUGUST 14, 2023 DR. JOANNA MELISSA F MARQUEZ, MD, MPH, DPSP _______________________________________________________________________________________________________ OBJECTIVES: 1. Review parts of the microscope...

ITM 101 | HISTOLOGY LESSON # 1 –INTRODUCTION TO HISTOLOGY 1st YEAR | A.Y. 2023-2024 | AUGUST 14, 2023 DR. JOANNA MELISSA F MARQUEZ, MD, MPH, DPSP _______________________________________________________________________________________________________ OBJECTIVES: 1. Review parts of the microscope. 2. Review types of microscopes. TYPES OF MICROSCOPES 3. Identification of cell and structures under the electron microscope. 4. Review cells and tissues under the microscope. 1. Bright field Simplest form of Light Microscopy light transmitted through or reflected from LIGHT MICROSCOPY it is collected to form an image that can be viewed. is used to make small structures and samples visible by Contrast and color in the image are formed providing a magnified image. because absorption and reflection vary over In microscopy, things that are invisible to the naked eye are seen the area of the sample. through a microscope. These are important for us to be able to Staining is necessary. Brightfield study physiological and pathological processes in the body. microscopy is the most common for all medical related courses. TWO (2) SUBSYSTEMS OF MICROSCOPE 2. Dark field Illumination System - The advantage is you the light source to illuminate the can see small structures that sample can’t be seen in light Imaging System - microscopy produces a magnified image of However, high the light that has interacted with illumination can destroy the the sample, which can then be specimen viewed by eye or using a camera system. 3. Phase Contrast (E.g. To see movement of larvae) Enables imaging of transparent PARTS OF MICROSCOPE samples 1. Eyepiece/Ocular Lens - (one Enables imaging of transparent (1) eyepiece - SIMPLE, two (2) eyepiece samples - COMPOUND) requires manipulation of both 2. Objective Turret, Revolver,or the illumination system and the Revolving Nosepiece - to hold multiple imaging system using apertures objective lenses. and filters; complex 3. Objective lenses, focus knobs - Used for tracking cell motion to move the stage. e.g. larva movement 4. Coarse adjustment - used in moving the stage higher or lower. 5. Fine adjustment - used to adjust 4. Polarized Light the focus of the lenses. This configuration 6. Stage - to hold specimen. requires a birefringement 7. Light source - a light or mirror. sample; example crystals. 8. Diaphragm and condenser - Strong background (ex: for lymphoma, the diaphragm/condenser or the amount of suppression from non- light that passes through is being adjusted to appreciate the birefringent areas of a sample, effacement of the lymph nodes) allows measurement of sample 9. Mechanical stage - has ruler that can measure the specimen. thickness and birefringence. Birefringent; will be encountered in concept of Gout (where birefringent sample is extracted at the knee) IMAGING SYSTEM Upper part of microscope. 5. Fluorescence Collects illuminating light that has Currently used at Ciudad Medical interacted with the sample and Center produces a magnified image that Allows individual fluorophores can be viewed. and particular areas of interest in This is achieved using two main a sample to be singled out groups of optical elements: o First, an objective lens that Can visualize the Cytoplasmic area or the Nucleus collects as much light from Important for Systemic Lupus the sample as possible; Erythematosus; Pattern of o Second, an eyepiece lens Fluorescence aids in diagnosis which relays the collected light to the observer’s eye or a camera system 101 INTRODUCTION TO MEDICINE | HISTOLOGY AYURA 2027 1 ITM 101 | HISTOLOGY LESSON # 1 –INTRODUCTION TO HISTOLOGY 1st YEAR | A.Y. 2023-2024 | AUGUST 14, 2023 DR. JOANNA MELISSA F MARQUEZ, MD, MPH, DPSP _______________________________________________________________________________________________________ 6. Electron Nucleus Electrons are used to illuminate the sample instead of visible The archive of the cell, the repository of its genetic material. light. Cell regulation, proliferation and DNA transcription Electrons have an associated wavelength which is much shorter The long DNA molecules contain the information necessary for than visible light, which allows magnifications of up to synthesis of all of the integral proteins and secretory products of 10,000,000. the cell. o DNA remains in the nucleus, it sends instructions to the cytoplasm via messenger RNA CELLS AND ORGANELLES o The synthetic activities of the cell are directed by informational macromolecules formed on the template of Used in studying the Organelles. DNA in the nucleus and transported to the cytoplasm to The cell is partitioned into two major compartments: Nucleus direct protein synthesis. and Cytoplasm Organelles; metabolically active internal organs - essential Inclusions; metabolically inert accumulations of cell products; dispensable Nucleus (Cont.) Centrally located, largest organelle of the cell The archive of the cell, the repository of its genetic material. In stained tissue, irregular clamps of chromatin may be seen Cell Membrane scattered throughout the nucleoplasm 8.5 to 10 nm in thickness Chromatin - composed of nucleic acids and histone proteins Contains local specializations for cell attachment and cell to cell that strongly bind hematoxylin and other basic dyes communication Principal nucleic acid of chromatin is deoxyribonucleic acid (DNA), which is stained with Feulgen reaction. o Euchromatin - extended state; sites of active transcription; light o Heterochromatin - condensed; inactive; dark Fluid mosaic model Main part of the cell membrane is the phospholipid bilayer. Phospholipids have a hydrophilic head and a hydrophobic tail. 101 INTRODUCTION TO MEDICINE | HISTOLOGY AYURA 2027 2 ITM 101 | HISTOLOGY LESSON # 1 –INTRODUCTION TO HISTOLOGY 1st YEAR | A.Y. 2023-2024 | AUGUST 14, 2023 DR. JOANNA MELISSA F MARQUEZ, MD, MPH, DPSP _______________________________________________________________________________________________________ Nuclear Envelope Golgi complex Nuclear envelope contains nuclear pores for some things to Control the traffic in small vesicles involved in the recycling of enter and exit (nucleoplasm within) membrane between organelles, and from the cytoplasm to the Controls the movement of macromolecules between the surface for renewal of the cell membrane. nucleoplasm and the surrounding cytoplasm. Electron microscope reveals it is composed of two parallel membranes separated by a 10-30 nm space , the perinuclear cisternae The Cytosol The cytosol is where metabolism, transport and protein translation occurs. Mitochondria Slender rods 0.4-0.8 um in diameter and 4-9 um in length. 2 membranes, one smooth outer membrane, and an inner membrane folded into layers called cristae Cristae has two compartments: the matrix and the intermembrane space Principal biochemical activity of mitochondria is oxidative phosphorylation. The energy generated is used to synthesize ATP. Hence, the powerhouse of the cell. Mitochondria divide before cell division, they are not synthesized like other cell parts Mitochondria possess their own DNA. It encodes the RNA of mitochondrial ribosomes. Rough endoplasmic reticulum Principal Biochemical Activity - Oxidative Phosphorylation Energy formed is stored as ATP - adenosine triphosphate Synthesis of membrane and export protein Contains ribosomes (20-25 nm particles) Ribosomes are sites where amino acids are assembled in the synthesis of proteins. With the aid of messenger RNA containing the information determining the sequence of the amino acids, polypeptides are formed. Smooth endoplasmic reticulum Protein modification, sorting and catabolism Involved in the synthesis of fatty acids and other lipids Found in abundance in steroid-secreting endocrine glands. In the liver, involved in VLDL synthesis 101 INTRODUCTION TO MEDICINE | HISTOLOGY AYURA 2027 3 ITM 101 | HISTOLOGY LESSON # 1 –INTRODUCTION TO HISTOLOGY 1st YEAR | A.Y. 2023-2024 | AUGUST 14, 2023 DR. JOANNA MELISSA F MARQUEZ, MD, MPH, DPSP _______________________________________________________________________________________________________ Lysosome Cytoskeleton Cellular catabolism Actin microfilaments - controls cell shape and movement Intracellular organelles containing degradative enzymes that Intermediate filaments. permit digestion of a wide range of macromolecules including o Rope-like structure/polymers, provides tensile strength protein, polysaccharide, lipids and nucleic acids. e.g., in epithelia intermediate filaments like desmosomes Defense against bacterial invasion and hemidesmosomes. Identification of acid phosphatase and other hydrolases is o Includes vimentin, desmin, neurofilaments, glial fibrillary important for verification of a lysosome organelle. acidic protein, cytokeratin, lamins. Microtubules o composed of alpha and beta tubulin dimers organized into hollow tubes. o regulated by the MTOC (microtubule organizing center), radiate from the centrosome o Also mediate sister chromatid segregation during mitosis ( forms the spindle during cell division) - Pulls during mitosis Toxic granulation is present when there is an increase in the number of large basophilic granules within circulating neutrophils. Levels of toxic granulation are highly correlated with C-reactive protein (CRP) values in patients with infection or inflammation. AN INTRODUCTION TO CELLS AND TISSUE SAMPLES IN LIGHT MICROSCOPY Peroxisome lacks hydrolase contains Urate oxidase, D-amino acid oxidase and catalase able to beta oxidate very long chain fatty acid producing H2O2 The image above showing Squamous cell or Squamous epithelial as byproduct cell from Pap Smear Clinical Importance: The patient is taking a drug → Normal: patterned, light colored, ample cytoplasm metabolized → produces many byproduct of hydrogen peroxide Attached blue dots: bacilli (may be from trichomonas vaginalis & infected) 101 INTRODUCTION TO MEDICINE | HISTOLOGY AYURA 2027 4 ITM 101 | HISTOLOGY LESSON # 1 –INTRODUCTION TO HISTOLOGY 1st YEAR | A.Y. 2023-2024 | AUGUST 14, 2023 DR. JOANNA MELISSA F MARQUEZ, MD, MPH, DPSP _______________________________________________________________________________________________________ The image above showing a Pathological Squamous cell from Pap Smear Abnormal: dark-colored, large malignant nucleus (actively The above image showing Large Cell Anaplastic Medublastoma, dividing), little cytoplasm, irregular the hematoxylin plays a key role in the diagnosis. Probably Cervical Cancer - Squamous Cell Carcinoma Black arrow: normalized acute inflammatory cell, similar in size to the RBCs on the previous image Note the mitotic figures (red arrow), indicating a rapidly growing tumor due to actively dividing cells. Infection from coccobacilli bacteria can lead to certain conditions in humans, such as bacterial vaginosis. The greater the number of mitosis, the greater the grade of carcinoma. Pattern Recognition - important in pathology, distinguish between Nuclear irregularity and chromatin clumping is also noted in the cells. normal and abnormal. Immunohistochemical Staining - used for cancers without pattern and looks different from others. In addition: cytoskeletal structures are used to find the origin of carcinoma, sarcoma or any malignancy that does not follow patterns. C E Image of a lymph node filled with Round Blue Cells. Hollow pink structures are the blood vessels. B D A The image above shows Skin H&E. Note the balanced coloration in this section of skin. The nuclei are stained purple, while the cytoplasmic components are pink. Layers of the Epidermis: (C LG Sobrang Bright) A. Stratum Corneum B. Stratum Lucideum C. Stratum Granulosum D. Stratum Spinosum E. Stratum Basale (Red Arrow) This is a stained image of the glomerulus of the kidney. The red arrows are pointing at Red blood cells (Erythrocytes) as they pass through the capillaries and exchange substances with the adjacent cells 101 INTRODUCTION TO MEDICINE | HISTOLOGY AYURA 2027 5 ITM 101 | HISTOLOGY LESSON # 1 –INTRODUCTION TO HISTOLOGY 1st YEAR | A.Y. 2023-2024 | AUGUST 14, 2023 DR. JOANNA MELISSA F MARQUEZ, MD, MPH, DPSP _______________________________________________________________________________________________________ The image shows a Colon H&E. Note the balanced coloration in this section of the colon. The microvilli on the columnar epithelium (Red Arrow) are very crisp. Glomerulus (red arrow) and tubules (blue arrow) displayed using H&E. The nuclei are stained purple, while the cytoplasmic components are pink. This image shows another part of the colon The red arrow is pointing at the mucin that is still visible with the goblet cells. 101 INTRODUCTION TO MEDICINE | HISTOLOGY AYURA 2027 6