MB 207 Molecular Cell Biology

1. MB 207 Molecular Cell Biology The lives and deaths of cells in tissues

2. The lives & Deaths of Cells in Tissues • Epidermis and its renewal by stem cells • Sensory epithelia • Blood vessels and epithelial cells • Renewal by pluripotent stem cells: Blood cell formation • Genesis, modulation and regeneration of skeletal muscle • Fibroblasts and their transformations: the connective tissue cell family • Stem cell engineering

3. Mammalian skin

4. Epidermal cells form a multilayered waterproof barrier

5. What is stem cells? • One of the body's master cells, with the ability to grow into any one of the body's more than 200 cell types • Unspecialized (undifferentiated) cells • Contribute to the body's ability to renew and repair its tissues. • Renew themselves as well as create new cells of whatever tissue they belong to (and other tissues). • The properties of a stem cells are: a) Not terminally differentiated b) can divide without limit (or at least for the life time of the animal) c) When it divides, each progeny has a choice – either remain a stem cell or cells that differentiate.

6. Stem cell biology • In the normal adult body, different classes of stem cells are responsible for the renewal of different types of tissue. • Some tissues are incapable of repair by the genesis of new cells because no competent stem cells are present. • Recent discoveries have opened up new possibilities for manipulating stem-cell behaviour artificially for repairing of un-repairable tissues. • Example: i) Epidermal stem cells taken from undamaged skin of a badly burnt patient can be rapidly grown in large numbers in culture and grafted back to reconstruct an epidermis to cover the burns. ii) Neural stem cells persist in a few regions of the adult mammalian brain and when grafted into a developing or damaged brain can generate new neurons and glia appropriate to the site of grafting. → Stem-cell biology offer hope of remedy for many serious diseases (eg. Parkinson disease).

7. Stem cell biology – Adult and Embryonic stem cells • Embryonic stem cells (ES cells) - from embryos (human or animal) - cells able to differentiate into many cell types in the body - can be induce to differentiate into different cell types in culture, when supplemented with certain growth/differentiation factors. • Adult stem cells - from some adult tissues (eg. bone marrow) - in a suitable environment, are able to generate a wider range of differentiated cell types than normally.

8. Stem cell division • Each daughter produced can either remain a stem cell or go on to become terminally differentiated. • Two ways in which stem cells produce daughters with different fate: i) based on environmental asymmetry - daughters of the stem cell are initially similar - are directed into different pathways according to the environmental influences that act on them - number of stem cells can be increased or reduced to fit the niche available for them ii) based on divisional asymmetry - the stem cell has an internal asymmetry - cell divides in such a way that its two daughters readily contained different determinants.

9. Two ways for a stem cell to produce daughters with different fates

10. Stem cell division • Stem cells in many tissues divide at a low rate. • Cell division give rise to transit amplifying daughters cells. • Transit cells are committed to differentiation - undergoes additional more rapid cell divisions to complete differentiation.

11. Transit Amplifying Cells In the example shown here, each stem cell division gives rise in this way to 8 terminally differentiated progeny

12. Role of Stem Cells • Stem cells division required when there is a need to replace differentiated cells (cannot divide) in a great variety of tissues. • Many types of stem cells, specialized for the genesis/generation of different classes of terminally differentiated cells Eg. Epidermal stem cells for epidermis Intestinal stem cells for intestinal epithelium Hemopoietic stem cells for blood etc… • Many factors determining whether stem cells divide or stay quiescent, progeny stay stem cells or differentiate = controlled by interactions with a variety of signals from surroundings or neighboring cells

13. What is Tissue Regeneration? • aims to develop living cell based biological approaches to aid the repair and regeneration of damaged and diseased tissues. • incorporates and links the areas of tissue engineering and regenerative medicine. • to develop living tissues, such as blood vessels, bone, cartilage, tendons, nerves in the laboratory, to implant into patients to recover physiological functions; to use synthetic and natural biomaterials to provide structure and form for laboratory assembled tissues and to use natural biological signals to direct tissue repair and to engage patient responses in completing the repair process. • is interdisciplinary research which combines the skills of biologists with material, engineering and physical scientists and with clinical and surgical expertise.

14. Tissue regeneration: Skin • The skin viewed as a large organ composed of 2 main tissues: the epidermis and the underlying connective tissue (consist of dermis & hypodermis). • Each tissue is composed of a variety of cell types. • The dermis and hypodermis are richly supplied with blood vessels and nerves (nerve fibers extend into the epidermis).

15. Regeneration of Skin • Consist of tough connective tissue (the dermis & hypodermis) overlaid by a multilayered waterproof epithelium (the epidermis). • Epidermis –continually renewed from stem cells (turnover time ~1 month in human). • Epidermal stem cells are clustered near the tips of the dermal papillae, attached to basal lamina. • Fate of these stem cells are controlled by interactionswith basal lamina and variety of signals from neighboring cells – regulate rate of basal cell proliferation in time of need. • Stem cell divide infrequently, giving rise (through a sideways movement) to transit amplifying cells. • The transit amplifying cells undergo several rapid divisions in the basal layer and then stop dividing, begin to differentiate and slip out of the basal layer. • They progressivelydifferentiateto form keratinocytes, switching from expression of one set of keratins to another until their nuclei degenerate, giving rise to an outer layer of dead keratinized cells that are continually shed from the surface. • Glands in the epidermis (mammary glands) have their own stem cells and their distinct patterns of cell turnover. - eg. Circulating hormones stimulate cells to proliferate and differentiate.

16. The distribution of stem cells in human epidermis, and the pattern of epidermal cell production.

17. Localization of different cells in skin regeneration: • Stem cells was identified by staining for b1-integrin. • Dividing cells were identified by labeling with BrdU (a thymidine analog that is incorporated into cells in S-phase of the cell division cycle). • Differentiating cells by staining for keratin-10 (a marker of keratinocyte differentiation).

18. Regeneration of Sensory receptor, Intestine and Liver Sensory receptor cells • eg. Epidermal cells and nerve cells = derived from the epithelium. • Olfactory receptor cells in the nose are full-fledge neurons. • They have a lifetime of 1-2 month and are continually replaced by new cells from stem cells in the olfactory epthelium. Intestine (Gut) • Expose to potentially damaging chemical processes = absorptive epithelium undergo constant & rapid renewal. • In small intestine, stem cells in the crypts generate new absorptive, goblet, enteroendocrine and Paneth cells, replacing most of the epithelial lining of the intestine every week. • The diverse fate of stem cell progeny are controlled by Notch signaling pathway, while the Wnt pathway is required to maintain the stem-cell population Liver • more protected organ, can rapidly adjust it’s size by cell proliferation or cell death when need arises. • differentiated hepatocytes = able to divide throughout life. • a specialized class of stem cells is not always needed for tissue renewal.

19. Sensory epithelia

20. The structure of the retina • photoreceptors are permanent cells that do not divide and are not replaced if destroyed. • photosensitive rhodopsin molecules are not permanent but are continually degraded and replaced.

21. Regeneration of blood vessels by endothelial cells • Endothelial cells form a single cell layer that lines all blood vessels and regulated exchanges between the blood stream and surrounding tissues. • Provide signals for organization and development of connective tissue cells that form the layers of blood-vessel wall. • New blood vessels can develop from walls of existing small vessels by the outgrowth of the endothelial cells. • Have the ability to form hollow capillary tubes in culture. • Expression of different cell-surface proteins - serve as control of proliferation. • In regions where cells are short of O2, increase in hypoxia-inducible factor 1 (HIF-1) stimulates the production of endothelial growth factor (VEGF) = causes endothelial cells to proliferate and invade the hypoxia tissue to supply new blood vessels.

22. Renewal by pluripotent (multipotent) stem cells: blood cell formation Pluripotent has the potential to differentiate into any of the three germ layers: endoderm, mesoderm, or ectoderm.

23. Types of white blood cells

24. Regeneration of blood cells • The different types of blood cells are all derived from a common multipotent stem cell (hemopoietic stem cells). • In adult, hemopoietic stem cells are in bone marrow and they depend on contact-mediated signals from the marrow stromal cells (connective tissue) to maintain their stem-cell character. • Hemopoietic stem cell normally divides infrequently to produce either multipotent stem cells (self-renewing) or committed progenitor cells (transit amplifying cells) each able to give rise to one or more types of blood cells. • Committed progenitor cells = lymphoid or the myeloid type. • The process where hemopoietic stem cells divide to form committed progenitor cells that differentiate to form blood cells is called hemopoiesis. • The committed progenitor cells divide extensively controlled by various protein signal molecules (colony-stimulating factors, CSFs) and subsequently terminally differentiated into mature blood cells having lifespan of several days or weeks.

25. Lymphoid progenitor cells give rise to: • T-cells (lymphocytes) develop in Thymus, • B-cells • NK cells • dendritic cells. • Myeloid progenitor cells give rise to: • monocytes (some dendritic cells,macrophages & osteoclasts), • granulocytes [neutrophils, eosinophils & basophils (devp Mast cells)] • megakaryocyte (platelets) • erythrocytes (RBCs). • Study of hemopoiesis = enhanced by in vitro assays in which stem cells or committed progenitor cells form clonal colonies when cultured in a semisolid matrix in the presence of CSF.

26. Rescue of an irradiated mouse by a transfusion of bone marrow cells

27. A tentative scheme of hemopoiesis

29. Genesis, modulation and regeneration of skeletal muscle • New skeletal muscle fibers form by the fusion of myoblasts • Muscle cells can vary their properties by changing the protein isoforms they contain • To control mucsle cell number and muscle cell size: loss of function mutation in myostatin, an extracellular TGFβ protein, can cause “double-muscled” animal. Both the numbers and the size of muscle cells seems to be increased.

30. Fibroblasts and their transformations: the connective tissue cell family Fibroblasts change their character in response to chemical signals. • When tissue is injured, the fibroblasts nearby proliferate, migrate into the wound and produce large amounts of collagenous matrix, which helps to isolate and repair the damaged tissue • The extracellular matrix may influence connective-tissue cell differentiation by affecting cell shape and attachment.