Las in vitro

Polona Tratnik

Las in vitro predstavlja del večletnega transdisciplinarnega raziskovalnega projekta, v sklopu katerega skupina sodelavcev goji lase v in vitro pogojih, za kar po pridobitvi človeškega vzorca tkiva z lasmi izvede vrsto eksperimentov, s katerimi dosega čim daljšo življenjsko dobo las v nadzorovanem, od telesa ločenem okolju, in živost tudi dokazuje, vizualizira. Za ta namen realizira sekvenčno slikanje gojenih las, ki prek fotografiranja rastočega materiala v času pokaže, kako se celice delijo in tvorijo lasno formo. Eksperimenti vključujejo eliminacijo matičnih celic iz vzorcev in preverjanje vpliva matičnih celic na gojišče z lasmi in pripadajočimi kožnimi celicami.

Projekt je rizomatsko strukturiran na več nivojih in spaja tehnoznanost z različnimi umetniškimi strategijami ter s humanističnim raziskovanjem tkivnega inženiringa in imunologije kot družbeno posebej aktualnih področij biotehnologije, ki obetata revolucionarne posledice še zlasti v medicini in estetski kirurgiji, ter refleksijo o hibridizaciji umetnosti, humanistike in tehnoznanosti, ki je v zadnjih letih v vzponu.

Avtorsko-izvajalska skupina je na različnih segmentih osredotočena na raziskovalni proces in spajanje področij ob usklajevanju heterogenih interesov. Pri tem delovanje ni usmerjeno v proizvajanje zaključenih proizvodov, artefaktov, namenjenih kontemplaciji, temveč v odpiranje raziskovalnega procesa in celotnega diskurza priložnostni javnosti. Namen projekta je komuniciranje biotehnoloških možnosti s širšo javnostjo, uresničevanje specifičnih ciljev, ki so zanimivi tako z biotehnološkega kot kiparskega in drugih aspektov, ter odpiranje povezanih vprašanj, ki so izrednega pomena za sodobnega posameznika in družbo.

»Medij, ki se uporablja v situaciji, kjer se v realnem času izvaja manipulacija z živim materialom in vitro, ni več medij, ki bi posredoval podobo, niti ni več medij, v katerem bi se življenje utelesilo, temveč je medij kot nadomestek telesnega okolja z vso njegovo vlažnostno, temperaturno in drugačno fragilnostjo, ki pogojno, če je eksperiment uspešen, omogoča celično preživetje za določen čas. Življenje, ki ga stalno ogroža smrt, je tu prisotno v izjemni občutljivosti in lahko vsak hip ugasne, pri tem pa njegovo prenehanje navzven ne bo prisotno kot meja, črta, lom, temveč je v njem že ves čas vsebovano kot odstavljeno za nedoločen čas vnaprej, a tudi za nazaj; prav tako pa je v prenehanju življenje prisotno kot možnost, s katero upravlja človek ob pomoči biotehnologije.«

»Umetniške situacije, ki gesto obvladovanja in nadzorovanja življenja ponavljajo, s tem ne ponavljajo tudi same družbene težnje po sistematičnem doseganju absolutnosti, temveč prej nasprotno: razprejo družbeno telo v njenem jedru, z vzpostavljanjem rizomatičnega telesa potopitveno (tukaj in zdaj, znotraj in zunaj) subjekt postavijo v brezorientacijsko okolje-telo, v katerem postane shizofrena celica, ki se shizofrenosti sebe-družbe v afektivnem soočenju tudi zave.«

»V planomenonih, planih oziroma ravneh konsistence ali kompozicije takšnega transdisciplinarnega hibrida, ki niso več hierarhično organizirane, se zgoščajo enkrat pretežneje biotehnološke, drugič umetniške, tretjič humanistične, četrtič še katere druge kolonije, ki pa nikoli niso očiščene drugih, elementi ene so postali sateliti druge. Takšne kolonije je postalo nemogoče ločevati med seboj, saj so med seboj prepredene in delujejo sočasno ter v medsebojnem usklajevanju.«

doc. dr. Polona Tratnik, In vitro.
Živo onstran telesa in umetnosti,
Koper: Univerza na Primorskem, 2010

»Da bi in vitro gojili lase in jih potem dejansko uporabljali v estetski kirurgiji – glede tega menim, da se mogoče celo nikoli ne bo zgodilo. A lasje v kulturah nam bodo omogočili bolje spoznati celotno fiziologijo las, njihov način razmnoževanja in tudi to, kako določene substance vplivajo na lase. To pomeni, da so lasje model za odkrivanje in preizkušanje novih zdravil. To je tudi namen naše raziskovalne naloge v sklopu projekta, ki ga podpira Javna agencija za raziskovalno dejavnost RS. Cilj, ki ga želimo izpeljati, je ugotoviti, kako lahko tudi celice iz lasnega mešička prispevajo k celotni skupini uporabnih matičnih celic. To pomeni nov vir celic, ki jih lahko potem uporabljamo tudi v drugih delih regenerativne medicine.«

»Sodelovanje me je pritegnilo predvsem zaradi opažanja, da obstaja velik prepad med sodobno znanostjo in vsakodnevnim življenjem. Vidim, da nekateri ljudje od znanosti preveč pričakujejo oziroma imajo v vsakem primeru neko popačeno sliko, medtem ko znanstveniki pogosto menijo, da morajo delovati v svojem ozkem okolju, kjer je zaprta tudi komunikacija, ki težko pride ven. Ko pa pride, pa je to ali s pretiranim navdušenjem, še pogosteje pa s kakšno pretirano afero. Ko se zgodi slednje, je škodo zelo težko popraviti. Z umetnostjo lahko znanost komunicira z ljudmi in dobi njihov odziv tudi na drugačen način, tj. na čustveni ravni, na ravni vtisov, reakcij, ki jih umetnik zazna veliko lažje in bolj senzitivno kot znanstvenik s svojimi eksperimentalnimi metodami.«

doc. dr. Miomir Knežević, univ. dipl. biol.,
Las in vitro, 15. junij 2010. © Horizonti.

»Naša zgodba presajanja in gojenja las morda ni neposredno uporabna, je pa zelo poučna. V vsakem takem projektu mora namreč sodelovati cela veriga strokovnjakov, potrebnih je veliko različnih znanj. Pri gojenju nekega preparata je kirurški del manjši, je v odvzemu in v nekem osnovnem razumevanju tkiva. Potem pa nastopi gojenje tkiva, določeni preparati itd. To so cele verige ljudi in znanj. Tisto, kar je zanimivo, je to sodelovanje in multidisciplinarnost, to, da hodiš preko neke svoje omejenosti, ki ji sicer lahko zelo hitro zapadeš, in pa to, da ustvariš nekaj novega in dobrega.«

»Umetnost se z resno znanostjo srečuje verjetno v tem, da izhajata iz istih korenov. Oboje izhaja iz nekega globokega človeškega iskanja. Ni le umetnost lepa, tudi prava znanost je lepa. Matematika je lepa. Ima neko transcendenco. Tako delo, igračkanje, raziskovanje, kot je v znanosti, je verjetno tudi v umetnosti. Tudi v umetnosti so ljudje, ki žrtvujejo življenje, ki so se pripravljeni odrekati, da nekam pridejo, da nekaj dosežejo, da se nekam potopijo. Mi pač nekaj imenujemo znanost, nekaj umetnost. Zdaj, ko je toliko možnosti, prihaja do prepleta. Najdejo se sorodne duše in se kaj rodi.«

Aleš Leskovšek, dr. med.,
spec. plast., rek., estet. kirurg.,
Las in vitro, 15. junij 2010. © Horizonti.

Citati

THE BIOLOGY OF HAIR FOLLICLES

»Hair has many useful biologic functions, including protection from the elements and dispersion of sweat-gland products (e.g., pheromones). It also has psychosocial importance in our society, and patients with hair loss (alopecia) or excessive hair growth often suffer tremendously.« p491

HAIR FOLLICULAR CELL/ORGAN CULTURE IN TISSUE ENGINEERING AND REGENERATIVE MEDICINE

»Hair follicles are complex organs composed of the dermal papilla (DP), dermal sheath (DS), outer root sheath (ORS), inner root sheath (IRS) and hair shaft. Development of hair follicles begins towards the end of the first trimester of pregnancy and is controlled by epidermal–mesenchymal interaction (EMI), which is a signaling cascade between epidermal and mesenchymal cell populations. Hair grows in cycles of various phases.« p323

»Conversely, the transplantation of human hair involves taking plugs of natural hair from areas in which occipital hair is growing and transplanting them to bald areas. However, the number of hairs that can be transplanted is limited in that only three such operations can generally be performed. To overcome such problems, many researchers have attempted to revive hair follicles by culturing hair follicle cells or mesenchymal cells in vitro and then implanting them in the treatment area.« p323

»A hair follicle is part of the skin that grows hair by packing old cells together. Sebaceous glands, which are tiny sebum-producing glands found everywhere except on the palms, lips and soles of the feet, are attached to each hair follicle. The thicker the density of hair, the more sebaceous glands are found. Stem cells are located at the junction of the sebaceous gland and the follicle and are primarily responsible for ongoing hair production via a process known as the anagen stage.« p324

»Hair follicles are divided into two major classes that occur in epidermal tissues and dermal tissues.« p324

»Cyclosporin A (CsA) has been used as a potent immunosuppressive agent for inhibition of graft rejection following organ transplantation.« p327

»Specifically, when hair follicles were cultured without insulin, they induced rapid transformation from anagen to catagen.« p327

»Substrates such as nylon mesh seeded with skin fibroblasts and full thickness skin were also found to support the growth of hair.« p327

»The evolution of hair follicle engineering began with the recognition in the early 1960s that hair follicles could be transplanted clinically into a foreign site and still grow a shaft typical of the donor site.« p329

SIMPLE AND RAPID METHOD TO ISOLATE AND CULTURE FOLLICULAR PAPILLAE FROM HUMAN SCALP HAIR FOLLICLES

»Published methodologies of HFP isolation rely on microdissecting manipulation for their successful isolation.« p381

»Moreover, imprecise manipulations at the level of the lower connective tissue sheath may increase the probability of dislodging and subsequently aspirating cells from the lower connective tissue sheath and/or epithelium. These cells may go on to contaminate the HFP culture.« p382

»Hair follicle papillae were individually isolated from intact isolated HF under bright-field stereoscopic illumination.« p382

»The quality of the HFP isolated using this method was assessed by high-resolution light microscopy (HRLM) and transmission electron microscopy.« p384

»Cultures of HFP are primarily obtained from terminal HFs.« p384

HUMAN HAIR GROWTH IN VITRO

»Human anagen hair follicles were isolated by microdissection from human scalp skin. Isolation of the hair follicles was achieved by cutting the follicle at the dermo-subcutaneous fat interface using a scalpel blade. Intact hair follicles were then removed from the fat using watchmakers' forceps. Isolated hair follicles maintained free-floating in supplemented Williams E medium in individual wells of 24-well multiwell plates showed a significant increase in length over 4 days.« p463

»Hair growth, which is effected by the division of the hair follicle matrix cells under control of the dermal papilla, is cyclical in the mammal.« p463

»These data show that in vitro hair follicles are able to maintain the in vivo pattern of DNA synthesis, and so it is reasonable to suppose that the production of a keratinised hair shaft in the maintained hair follicle occurs as a result of matrix cell division in the hair follicle bulb.« p468

HUMAN HAIR CYCLE

»The length of hair cycles in man varies in the different regions of the body.« p65

»Methods of measuring human hair cycles: calculation of the length of the cycle by measuring the final length of hair; marking the tips of individual hairs with ink dye; plucking hairs and observing their roots to determine the cycle periods.« p65

HAIR IN FORENSIC MEDICINE

»Possible applications of hair analysis include diagnosis of drug abuse, poisoning and doping, evaluation of occupational exposure to toxics, evaluation of prenatal exposure to drugs, and monitoring of a patient's compliance with drug prescription.« p539

»Drugs and toxics are incorporated into hair through three different modalities:
Passive diffusion to the hair matrix from the blood and successive incorporation in the hair shaft during keratinization. Transfer to the formed hair shaft from sebum and sweat. Transfer to the formed shaft from the environment.« p540

»Hair sampling is usually made from the vertex or from the pubis where hair is less contaminated by environmental and cosmetic factors.« p541

HAIR GROWTH ASSESSMENT TECHNIQUES

»Qualitative and quantitative methods are essential for objectively evaluating hair growth activity.« p125

CYCLOSPORIN A-INDUCED HAIR GROWTH IN MICE IS ASSOCIATED WITH INHIBITION OF CALCINEURIN-DEPENDENT ACTIVATION OF NFAT IN FOLLICULAR KERATINOCYTES

»Immunosuppressive immunophilin ligands such as cyclosporin A (CsA) and FK506 are known as potent hair growth modulatory agents in rodents and humans that induce active hair growth (anagen) and inhibit hair follicle regression (catagen).« p1593

»The immunosuppressive effectiveness of these drugs has been generally attributed to inhibition of T cell activation through well-characterized pathways.« p1593

»Growth and development of hair follicles are influenced by a variety of growth factors and cytokines, the most widely described being keratinocyte growth factor (KGF), interleukin-1 (IL-1), and transforming growth factor _ (TGF_).« p1594

AT THE ROOTS OF A NEVER-ENDING CYCLE

»Wnt pathways in the development of many different organs in the body, including limbs, cartilage, kidney, and brain (reviewed by Cadigan and Nusse, 1997).« p18

»While normal dermal papilla cells rapidly lose their inductive powers in culture (Jahoda et al., 1984), the Wnt-bathed dermal papilla cells maintained inductive capacity to generate hair follicles when trans- are planted back into mice.« p19

»A discussion of the biochemical events involved in Wnt signaling and hair follicle formation would not be complete without further discussion of the Shh signal transduction pathway.« p22

ANIMAL MODELS AND CULTURE METHODS IN THE STUDY OF HAIR GROWTH

»Further, if each follicle contains the clock signal, then a growth wave must arise from coordination of individual follicles by an overriding signal, presumably of endocrine origin.« p105

»In a recent discussion of hair cycling,2 an inhibitor theory was proposed in which the removal or release of an inhibitory molecule in the epithelium stimulates anagen and growth; however, this leads to the question as to what is the nature of the event that removes the inhibitor.« p105

»Culture systems should become increasingly important in the dissection of follicle cell function in hair diseases.« p110

»The important interactions known to occur between the epithelial and mesenchymal components of skin can best be studied using in vitro models for all of the above reasons; however, there are also some potential disadvantages of in vitro models that should not be ignored.« p110

CHARACTERIZATION OF A NEW TISSUE-ENGINEERED HUMAN SKIN EQUIVALENT WITH HAIR

»This model was produced exclusively from human fibroblasts and keratinocytes and did not contain any synthetic material.« p318

»Therefore, this new hairy, human skin equivalent mode/allowed an experimental design in which the only variable was the presence of pilosebaceous units and provided new data confirming the importance of hair follicles in pereutaneous absorption.« p318

»Hair follicles have been previously cultured in vitro, in an organ culture system, or in sponge-supported histoculture (Philpott et al., 1990; Li et al. 1992).« p318

»Hair follicles have also been used as a source of keratinoeytes in a skin equivalent model (Lenoir et al., 1988; Lenoir-Viale et al., 1993).« p318

»Hairless animals are not reliable models because they still have hair follicles.« p319

»Human keratinocytes and dermal fibroblasts were obtained from normal adult skin specimens removed during reductive breast surgery.« p319

»Human fibroblasts were obtained from the dermal portion of skin biopsies.« p319

»Control samples (skin equivalents with sham hair insertion) were obtained by laying down an explant of interfollicular epidermis (i.e., without hair follicle) on top of the holes previously made in the dermal equivalent. An additional fibroblast sheet was also added under the dermal equivalent.« p319

»For histological analysis, skin equivalents were fixed with Bouin's solution and paraffin embedded. Six-lam-thick sections were stained with hematoxylin, phloxine, and saffron.« p319

THE BIOLOGY OF HAIR GROWTH

»Hair follicles go through well established repeated cycles of development and growth (anagen), regression (catagen), and rest (telogen) to enable the replacement of hairs, often by another of differing colour or size. An additional phase, exogen, has been reported where the resting club hair is released.« p10

1. Green M.R., Clay C.S., Gibson W.T., Hughes T.C., Smith C.G., Westgate G.E., White M. in Kealey T. 1986. Rapid Isolation in Large Numbers of Intact, Viable, Individual Hair Follicles From Skin: Biochemical and Ultrastructural Characterization. J Invest Dermatol, 87:768-770

2. The strip of skin was repetedly cut with a loose-fitting pair pf scissors for 1-3 m and the supernatant liquid removed and examined under Wild M8 binocular dissecting microscope. Str.768

3. Keug P. J., Hoon L.D., Hoon Y.H., Ho S.Y., Yong S.K. 2006. Method for the preparation of a dermal papilla tissue having hair follicle inductive potency. KR20040098680 20041129 – PATENT Živa

4. Philpott M.P., Green M.R. in Kealey T. 1989. Studies on the biochemistry and morphology of freshly isolated and maintained rat hair follicles. Journal of Cell Scienc 93, 409-418
We now show that these follicles can be viably maintained on permeable supports for 7 days in vitro as determined by their adenine nucleotide contents, rates of [mefliy/-3H]thymidine and [U-14C]leucine uptake, [methyl- H]thymidine autoradiography, patterns of keratin synthesis and light and electron microscopy. Str. 409

5. Philpott M.P., Green M.R. in Kealey T. 1990. Human hair growth in vitro. Journal of Cell Scienc 97, 463-471
We report for the first time the successful maintenance and growth of human hair follicles in vitro.
Isolated hair follicles maintained free-floating in supplemented Williams E medium in individual wells of 24-well multiwell plates showed a significant increase in length over 4 days. Str. 463

Richardson G.D., Arnott E. C., Whitehouse C. J., Lawrence C. M., Hole N., in Jahoda C.A.B. 2005. Cultured Cells from the Adult Human Hair Follicle Dermis can be Directed Toward Adipogenic and Osteogenic Differentiation. J Invest Dermatol, 124:1090-1091
Here, we report adipogenic and osteogenic differentiation in human hair follicle-derived DP and DS cultures. Str 1090

Kim1 J.Y.,, Tavaré S. in Shibata D. 2006. Human hair genealogies and stem cell latency. BMC Biology 2006, 4:2
Average hair errors increased until two years of age, and then were constant despite decades of replacement, consistent with new hairs arising from infrequently dividing bulge stem cells. Errors were significantly more frequent in longer hairs, consistent with long-lived but eventually shed mitotic follicle cells. Str. 1

Lyle S., Christofidou-Solomidou M., Liu Y., Elder1 D.E., Albelda S. in Cotsarelis G. 1998. The C8/144B monoclonal antibody recognizes cytokeratin 15 and defines the location of human hair follicle stem cells. Journal of Cell Science 111, 3179-3188
The homeostasis of all self-renewing tissues, including the epidermis and hair follicle, is thought to be dependent on stem cells As undifferentiated stem cells divide, they generate daughter cells that retain the stem cell phenotype, and daughter cells (called transit-amplifying (TA) cells) that undergo rapid proliferation and terminal differentiation to repopulate the tissue str. 3179

Porter R.M. 2003. REVIEW: Mouse models for human hair loss disorders. J. Anat. 202, 125–131
Changes in hair growth str. 127

Spradling A., Drummond-Barbosa D. in Toshie Kai. 2001. insight review articles: Stem cells find their niche. Nature vol. 414., 98-104
Hair follicles contain two zones of stem-like cells. The hair shaft and its surrounding sheaths are produced by matrix cells located on a basement membrane overlying the protruding piece of dermis know as the dermal papilla. Str. 101

Jones C.J. in Kealey T. 1987. Electrophysiological and dye-coupling studies on secretory, myoepithelial and duct cells in human eccrine sweat glands. J.Physiol. 389, 461-481

LEE C.M., JONES C.J. in Kealey T. 1984. Biochemical and ultrastructural studies of human eccrine sweat glands isolated by shearing and maintained for seven days. J.Cell Sci. 72, 259-274
Sweat glands were picked out using two pairs of stainless steel Dumont no. 5 microforceps. Str. 206

Članki

PRAVNI AKTI / LEGAL ACTS:

Človekove pravice v zvezi z biomedicino: Oviedska konvencija in dodatni protokoli. 2009. Ministrstvo za zdravje RS, Ljubljana, 104.

Biomedicine and human rights; The Oviedo Convention and its additional protocols. 2009. Council of Europe, 190.

Kodeks etike zdravstvenih delavcev Slovenije. 2002. Ur.l. RS št. 4/2002, 406.

Konvencije Sveta Evrope o varovanju človekovih pravic in dostojanstva človeškega bitja. 1998. Ur.l. RS MP 17/98, 277.

Convention for the Protection of Human Rights and Dignity of the Human Being with regard to the Application of Biology and Medicine: Convention on Human Rights and Biomedicine. 1997. Council of Europe, 20.

Načela Helsinške deklaracije o biomedicinskih raziskavah na človeku. 2008. 59th WMA General Assembly, Seoul.

Declaration of Helsinki - Ethical Principles for Medical Research Involving Human Subjects. 2008. World Medical Association.


VIRI / SOURCES:

»The length of hair cycles in man varies in the different regions of the body.« 65
»Methods of measuring human hair cycles: calculation of the length of the cycle by measuring the final length of hair; marking the tips of individual hairs with ink dye; plucking hairs and observing their roots to determine the cycle periods.« 65

Vir/Source: Alonso L., Fuchs E. 2006. The hair cycle. Journal of Cell Science, 119, 391–393.

»Qualitative and quantitative methods are essential for objectively evaluating hair growth activity.« 125

Vir/Source: Blume-Peytavi U., Hillmann K., Guarrera M. 2008. Hair growth assessment techniques. Hair growth and disorders. Whiting D. A., Trϋeb R. M. (eds.). Leipzig, Springer, 125–157.

»Wnt pathways in the development of many different organs in the body, including limbs, cartilage, kidney, and brain (reviewed by Cadigan and Nusse, 1997).« 18
»While normal dermal papilla cells rapidly lose their inductive powers in culture (Jahoda et al., 1984), the Wnt-bathed dermal papilla cells maintained inductive capacity to generate hair follicles when trans- are planted back into mice.« 19

»A discussion of the biochemical events involved in Wnt signaling and hair follicle formation would not be complete without further discussion of the Shh signal transduction pathway.« 22

Vir/Source: Fuchs E., Merrill B.J., Jamora C., DasGupta R. 2001. At the roots of a never-ending cycle. Developmental cell, 1, 13–25.

»Immunosuppressive immunophilin ligands such as cyclosporin A (CsA) and FK506 are known as potent hair growth modulatory agents in rodents and humans that induce active hair growth (anagen) and inhibit hair follicle regression (catagen).« 1593
»The immunosuppressive effectiveness of these drugs has been generally attributed to inhibition of T cell activation through well-characterized pathways.« 1593
»Growth and development of hair follicles are influenced by a variety of growth factors and cytokines, the most widely described being keratinocyte growth factor (KGF), interleukin-1 (IL-1), and transforming growth factor _ (TGF_).« 1594

Vir/Source: Gafter-Gvili A., Sredni B., Gal R., Gafter U., Kalechman Y. 2003. Cyclosporin A-induced hair growth in mice is associated with inhibition of calcineurin-dependent activation of NFAT in follicular keratinocytes. AJC Cell Physiology, 284, 1593–1603.


»The strip of skin was repetedly cut with a loose-fitting pair pf scissors for 1-3 m and the supernatant liquid removed and examined under Wild M8 binocular dissecting microscope.« 768

Vir/Source: Green M.R., Clay C. S., Gibson W. T., Hughes T. C., Smith C. G., Westgate G. E., White M., Kealey T. 1986. Rapid Isolation in Large Numbers of Intact, Viable, Individual Hair Follicles From Skin: Biochemical and Ultrastructural Characterization. J Invest Dermatol, 87, 768–770.

»The strip of skin was repetedly cut with a loose-fitting pair pf scissors for 1-3 m and the supernatant liquid removed and examined under Wild M8 binocular dissecting microscope.« 768

Vir/Source: Green M. R. et. al. 1986. Rapid isolation in large numbers on intact, viable, individual hair follicles from skin: Biochemical and ultrastructural characterization. J. Invest Dermatol 87, 768–770.

»Average hair errors increased until two years of age, and then were constant despite decades of replacement, consistent with new hairs arising from infrequently dividing bulge stem cells. Errors were significantly more frequent in longer hairs, consistent with long-lived but eventually shed mitotic follicle cells.« 1

Vir/Source: Kim1 J. Y.,, Tavaré S. in Shibata D. 2006. Human hair genealogies and stem cell latency. BMC Biology 2006, 4, 2.

»Sweat glands were picked out using two pairs of stainless steel Dumont no. 5 microforceps.« 206

Vir/Source: Lee C. M., Jones C. J., Kealey T. 1984. Biochemical and ultrastructural studies of human eccrine sweat glands isolated by shearing and maintained for seven days. J.Cell Sci. 72, 259–274.

»The homeostasis of all self-renewing tissues, including the epidermis and hair follicle, is thought to be dependent on stem cells As undifferentiated stem cells divide, they generate daughter cells that retain the stem cell phenotype, and daughter cells (called transit-amplifying (TA) cells) that undergo rapid proliferation and terminal differentiation to repopulate the tissue.« 3179

Vir/Source: Lyle S., Christofidou-Solomidou M., Liu Y., Elder1 D.E., Albelda S. in Cotsarelis G. 1998. The C8/144B monoclonal antibody recognizes cytokeratin 15 and defines the location of human hair follicle stem cells. Journal of Cell Science 111, 3179–3188.

»Published methodologies of HFP isolation rely on microdissecting manipulation for their successful isolation.« 381
»Moreover, imprecise manipulations at the level of the lower connective tissue sheath may increase the probability of dislodging and subsequently aspirating cells from the lower connective tissue sheath and/or epithelium. These cells may go on to contaminate the HFP culture.« 382
»Hair follicle papillae were individually isolated from intact isolated HF under bright-field stereoscopic illumination.« 382
»The quality of the HFP isolated using this method was assessed by high-resolution light microscopy (HRLM) and transmission electron microscopy.« 384
»Cultures of HFP are primarily obtained from terminal HFs.« 384

Vir/Source: Magerls M., Kauser S., Tobin P. R. 2002. Simple and rapid method to isolate and culture follicular papillae from human scalp hair follicles. Experimental Dermatology, 11, 381–385.

»This model was produced exclusively from human fibroblasts and keratinocytes and did not contain any synthetic material.« 318
»Therefore, this new hairy, human skin equivalent mode/allowed an experimental design in which the only variable was the presence of pilosebaceous units and provided new data confirming the importance of hair follicles in pereutaneous absorption.« 318
»Hair follicles have been previously cultured in vitro, in an organ culture system, or in sponge-supported histoculture (Philpott et al., 1990; Li et al. 1992).« 318
»Hair follicles have also been used as a source of keratinoeytes in a skin equivalent model (Lenoir et al., 1988; Lenoir-Viale et al., 1993).« 318
»Hairless animals are not reliable models because they still have hair follicles.« 319
»Human keratinocytes and dermal fibroblasts were obtained from normal adult skin specimens removed during reductive breast surgery.« 319
»Human fibroblasts were obtained from the dermal portion of skin biopsies.« 319
»Control samples (skin equivalents with sham hair insertion) were obtained by laying down an explant of interfollicular epidermis (i.e., without hair follicle) on top of the holes previously made in the dermal equivalent. An additional fibroblast sheet was also added under the dermal equivalent.« 319
»For histological analysis, skin equivalents were fixed with Bouin's solution and paraffin embedded. Six-lam-thick sections were stained with hematoxylin, phloxine, and saffron.« 319

Vir/Source: Michel M., L'heureux N., Pouliot R., Xu W., Auger F. A., Germain L. 1999. Characterization of a new tissue-engineered human skin equivalent with skin. In Vitro Cellular & Developmental Biology-Animal, 35, 6, 318–326.

»Hair has many useful biologic functions, including protection from the elements and dispersion of sweat-gland products (e.g., pheromones). It also has psychosocial importance in our society, and patients with hair loss (alopecia) or excessive hair growth often suffer tremendously.« 491

Vir/Source: Paus F., Cotsarelis G. 1999. Mechanisms of disease: The biology of hair follicles. The New England Journal of Medicine, 341, 7, 491–497.

»We report for the first time the successful maintenance and growth of human hair follicles in vitro. Isolated hair follicles maintained free-floating in supplemented Williams E medium in individual wells of 24-well multiwell plates showed a significant increase in length over 4 days.« 463
»Human anagen hair follicles were isolated by microdissection from human scalp skin. Isolation of the hair follicles was achieved by cutting the follicle at the dermo-subcutaneous fat interface using a scalpel blade. Intact hair follicles were then removed from the fat using watchmakers' forceps. Isolated hair follicles maintained free-floating in supplemented Williams E medium in individual wells of 24-well multiwell plates showed a significant increase in length over 4 days.« 463
»Hair growth, which is effected by the division of the hair follicle matrix cells under control of the dermal papilla, is cyclical in the mammal.« 463
»These data show that in vitro hair follicles are able to maintain the in vivo pattern of DNA synthesis, and so it is reasonable to suppose that the production of a keratinised hair shaft in the maintained hair follicle occurs as a result of matrix cell division in the hair follicle bulb.« 468

Vir/Source: Philpott M., Green M. R., Kealey T. 1990. Human hair growth in vitro. Journal of Cell Science, 97, 463–471.

»We now show that these follicles can be viably maintained on permeable supports for 7 days in vitro as determined by their adenine nucleotide contents, rates of [mefliy/-3H]thymidine and [U-14C]leucine uptake, [methyl- H]thymidine autoradiography, patterns of keratin synthesis and light and electron microscopy.« 409

Vir/Source: Philpott M. P., Green M. R. in Kealey T. 1989. Studies on the biochemistry and morphology of freshly isolated and maintained rat hair follicles. Journal of Cell Science 93, 409–418.

»(Changes in hair growth:) However, in humans, by far the most common cause of hair disorders are defects in hair growth control.« 127

Vir/Source: Porter R. M. 2003. REVIEW: Mouse models for human hair loss disorders. J. Anat. 202, 125–131.

»Possible applications of hair analysis include diagnosis of drug abuse, poisoning and doping, evaluation of occupational exposure to toxics, evaluation of prenatal exposure to drugs, and monitoring of a patient's compliance with drug prescription.« 539
»Drugs and toxics are incorporated into hair through three different modalities:
- Passive diffusion to the hair matrix from the blood and successive incorporation in the hair shaft during keratinization.
- Transfer to the formed hair shaft from sebum and sweat.
- Transfer to the formed shaft from the environment.« 540
»Hair sampling is usually made from the vertex or from the pubis where hair is less contaminated by environmental and cosmetic factors.« 541

Vir/Source: Piraccini B. M., Pazzaglia M., Tosti A. 2008. Hair in forensic medicine. Hair growth and disorders. Whiting D.A., Trϋeb R.M. (eds.). Leipzig, Springer, 539–542.

»Hair follicles go through well established repeated cycles of development and growth (anagen), regression (catagen), and rest (telogen) to enable the replacement of hairs, often by another of differing colour or size. An additional phase, exogen, has been reported where the resting club hair is released.« 10

Vir/Source: Randall V. A., Botchkareva N. V. 2009. The biology of hair growth. Cosmetic applications of laser and light-based systems. UK, William Andrew Inc., 3–35.

»Further, if each follicle contains the clock signal, then a growth wave must arise from coordination of individual follicles by an overriding signal, presumably of endocrine origin.« 105
»In a recent discussion of hair cycling,2 an inhibitor theory was proposed in which the removal or release of an inhibitory molecule in the epithelium stimulates anagen and growth; however, this leads to the question as to what is the nature of the event that removes the inhibitor.« 105
»Culture systems should become increasingly important in the dissection of follicle cell function in hair diseases.« 110
»The important interactions known to occur between the epithelial and mesenchymal components of skin can best be studied using in vitro models for all of the above reasons; however, there are also some potential disadvantages of in vitro models that should not be ignored.« 110

Vir/Source: Rogers G., Hynd P. 2001. Animal models and culture methods in the study of hair growth. Clinics in Dermatology, 19, 105–119.

»Here, we report adipogenic and osteogenic differentiation in human hair follicle-derived DP and DS cultures.« 1090

Vir/Source: Richardson G. D., Arnott E. C., Whitehouse C. J., Lawrence C. M., Hole N., in Jahoda C. A. B. 2005. Cultured Cells from the Adult Human Hair Follicle Dermis can be Directed Toward Adipogenic and Osteogenic Differentiation. J Invest Dermatol, 124, 1090–1091.

»Hair follicles contain two zones of stem-like cells. The hair shaft and its surrounding sheaths are produced by matrix cells located on a basement membrane overlying the protruding piece of dermis know as the dermal papilla.« 101

Vir/Source: Spradling A., Drummond-Barbosa D. in Toshie Kai. 2001. insight review articles: Stem cells find their niche. Nature vol. 414., 98–104.

»Hair follicles are complex organs composed of the dermal papilla (DP), dermal sheath (DS), outer root sheath (ORS), inner root sheath (IRS) and hair shaft. Development of hair follicles begins towards the end of the first trimester of pregnancy and is controlled by epidermal–mesenchymal interaction (EMI), which is a signaling cascade between epidermal and mesenchymal cell populations. Hair grows in cycles of various phases.« 323
»Conversely, the transplantation of human hair involves taking plugs of natural hair from areas in which occipital hair is growing and transplanting them to bald areas. However, the number of hairs that can be transplanted is limited in that only three such operations can generally be performed. To overcome such problems, many researchers have attempted to revive hair follicles by culturing hair follicle cells or mesenchymal cells in vitro and then implanting them in the treatment area.« 323
»A hair follicle is part of the skin that grows hair by packing old cells together. Sebaceous glands, which are tiny sebum-producing glands found everywhere except on the palms, lips and soles of the feet, are attached to each hair follicle. The thicker the density of hair, the more sebaceous glands are found. Stem cells are located at the junction of the sebaceous gland and the follicle and are primarily responsible for ongoing hair production via a process known as the anagen stage.« 324
»Hair follicles are divided into two major classes that occur in epidermal tissues and dermal tissues.« 324
»Cyclosporin A (CsA) has been used as a potent immunosuppressive agent for inhibition of graft rejection following organ transplantation.« 327
»Specifically, when hair follicles were cultured without insulin, they induced rapid transformation from anagen to catagen.« 327
»Substrates such as nylon mesh seeded with skin fibroblasts and full thickness skin were also found to support the growth of hair.« 327
»The evolution of hair follicle engineering began with the recognition in the early 1960s that hair follicles could be transplanted clinically into a foreign site and still grow a shaft typical of the donor site.« 329

Vir/Source: Yoo B., Shin Y., Yoon H., Seo Y., Park J. 2009. Hair follicular cell/organ culture in tissue engineering and regenerative medicine. Biochemical Engineering Journal, 48, 323–331.

Drugi viri / Other sources:

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