• 한국생물공학회:학술대회논문집
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• 2003.04a
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• pp.302-305
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• 2003

It is difficult to obtain sufficient healthy skin for coverage of a wide area of skin wound. In the skin, an additional population of living epithelial cells is located in the outer root sheath (ORS) of hair $follicles.^{1),2)}$ ORS cells should be a good source of epithelium because they are easily obtainable and patients do not have to suffer from scar formation at donor sites. We modified ordinary primary culture technique for the purpose of solving such problem that epithelial cells have a low propagation and easy aging during culture periods. First of all, we improved primary cultivation methods. In the ordinary primary culture, average yield of human ORS cells was $2\;{\times}\;10^3$ cells/follicle by direct incubation with trypsin (0.1%)/EDTA (0.02%) solution for 15 min at $37^{\circ}C$ but we could obtain about $6.5\;{\times}\;10^3$ cells/follicle by two step enzyme digestion method with dispase (1.2 U/ml) and trypsin (0.1%)/EDTA (0.02%) solution. So we could achieve three times higher primary cultured ORS cell yield. Secondly, we could obtain total $2\;{\times}\;10^7$ cells in serum free medium and even more total $6\;{\times}\;10^7$ cells in modified E-medium with mitomycin C-treated feeder cells during 17 days. Using the cultured ORS cells, and we could make bioartificial skin equivalent in vitro and concluded that ORS cells were progenitor cells for skin epithelial cell.

• The Journal of Korean Medicine
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• v.18 no.2
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• pp.273-282
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• 1997

To observe the effect of Heulbuchuketang on endometriosis, surgically induced endometriosis in rats was treated with Heulbuchuketang for l00days. The result are as follows: 1. Fragments of right uterine horn were implanted at the serosal wall of small intestine and its adjacent aterial suppling branch. After 4 weeks later endometriosis was observed at the area of implantation of small intestine. 2. After l00-day-treatment, the size of the endometriosis of treated group was much smaller compared to those of the untreated group. Especially the result of the later 60-day-treatment was much better than that of the early 40-day-treatment. 3. In case of inflammation or pustula induced from endometriosis, the size of abnormal endometriosis tissue showed no change after the total treatment and fibrotic change was often founded there, which was not responding to Heulbuchuketang. 4. In the ovary of untreated group, primordial follicle, primary follicle, maturing primary follicle were easily founded and there was no difference in shape and size between the ovary of Heulbuchuketang treated group. From the upper 1 to 4 laboratory result it can be suggested that Heulbuchuketang can be specially effective to mild endometriosis with conservating the ability of fertility.

• Journal of Microbiology and Biotechnology
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• v.34 no.3
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• pp.506-515
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• 2024

Primary human dermal papilla cells (HDPCs) are often preferred in studies on hair growth and regeneration. However, primary HDPCs are limited by their reduced proliferative capacity, decreased hair induction potential, and extended doubling times at higher passages. To overcome these limitations, pTARGET vectors containing human papillomavirus16 (HPV16) E6/E7 oncogenes were transfected into HDPCs and selected using G-148 to generate immortalized cells here. HPV16 E6/E7 oncogenes were efficiently transfected into primary HDPCs. Immortalized HDPC showed higher proliferative activity than primary HDPC, confirming an increased proliferation rate. Expression of p53 and pRb proteins was downregulated by E6 and E7, respectively. E6/E7 expressing HDPC cells revealed that cyclin-dependent kinase (CDK) inhibitor p21 expression was decreased, while cell cycle-related genes and proteins (CDK2 and cyclin E) and E2F family genes were upregulated. Immortalized HDPCs maintained their responsiveness to Wnt/β-catenin pathway and hair follicle formation capability, as indicated by their aggregative properties and stemness. E6/E7 immortalized HDPCs may facilitate in vitro hair growth and regeneration studies.

• The Korean Journal of Zoology
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• v.39 no.4
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• pp.410-418
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• 1996

Gangilosides are ubiquitous membrane components in mammalian cells and are suggested to play essential roles in cellular phenomena such as cell-cell interadion, differentiation, and signal transduction. Rat ovary contained GM3 as major gangiloside. Nn order to study GM3 distribution in the atretic follicles and its possible changes during follicular development, frozen sections were stained with spedfic monocional antibodies against eleven gangilo-series gangliosides including GM3. In the atretic follicles, Glf3 was expressed in a spatlo-temporally different manner during foilicular development, but GM1 and other gangliosides were not immunohistochemicaily detected. in atretic follicle from primary follicle stage, GM3 was expressed in all the theca cells and some granulosa cells adjacent to oocyte. In atretic follicle from secondary follicle stage, GM3 were expressed in all theca cells and granulosa cells. in atretic follicles from developing Graaflan follicle stages, GM3 was similarly expressed as in secondary follicle stage.

• Journal of Animal Reproduction and Biotechnology
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• v.34 no.1
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• pp.35-39
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• 2019

This study aimed to recover the ovarian function through in vitro culture of preantral follicles from aged mice. First, we isolated the preantral follicles from ovaries of sixty-seven-week old B6D2F1 mice with decreased fecundity to know how many follicles were present in them, which was 6 preantral follicles including 2 primary, 2 early secondary and late secondary follicles from 8 aged mice. It was confirmed that a few follicles (~2) were present in aged mice through histological analysis compared to adult mice as control. The 9 days of in vitro culture of preantal follicles showed in vitro growth and induced maturation after treatment with hCG (2.5 IU/mL) and EGF (5 ng/mL). Cumulus cells in the cumulus-oocyte complexes (COCs) were removed using hyaluronidase and oocytes at the germinal vesicle (GV) and GV breakdown (GVBD) were obtained from preantral follicle culture of aged mice in vitro. In conclusion, these observations demonstrated that there still were a few preantral follicles in the ovaries of 67 week-old mice, which we were able to culture in vitro and oocytes were obtained from them. This study proposed an in vitro culture system using preantral follicle as a therapeutic strategy for fertility preservation in humans for assisted reproductive medicine.

• Animal Bioscience
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• v.34 no.9
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• pp.1544-1551
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• 2021

Objective: The purpose of this study was to investigate growth performance, carcass traits, meat quality and hair follicle development of growing Rex rabbits as affected by different environmental enrichment materials. Methods: A total of one hundred and twenty Rex rabbits were randomly assigned to four groups; reared in conventional cages (not enriched) and in enriched cages with either willow stick (WS), rubber duck, or a can containing beans (CB), for 44 days. Results: The average daily gain of the CB group was the highest and had a significant difference from that of the other groups (p<0.05). The spleen and cecum weight of the CB group was greater than those of the WS and control groups (p<0.05). The redness (Commission Internationale de l'Eclairage a^*) of the meat sample of the control group was lower than those of the enriched cage groups (p<0.05). Moreover, the hue value of the CB group was significantly lower than that of the other groups (p<0.05). The tenderest meat belonged to the CB group. In addition, more secondary (p<0.05) and primary follicles were found in the CB group than in the control group. Conclusion: Environmental enrichment increased the average daily gain and improved some carcass traits, meat quality, and hair follicle density. Among the three environmental enrichment materials, CB could be recommended for rabbit husbandry.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.11
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• pp.1761-1769
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• 2007

This experiment was conducted to determine the extent to which susceptibility to cortisol-induced follicle shutdown is influenced by fleece phentotype. Twenty Finewool (10 sheep low fibre diameter, low coefficient of fibre diameter-LL and 10 low fibre diameter, high coefficient of variation of fibre diameter-LH) and twenty Strongwool (10 low fibre diameter, low coefficient of variation of fibre diameter-HL and 10 high fibre diameter and high coefficient of variation of fibre diameter-HH) sheep of 9 months of age were individually penned in an animal house and were injected intramuscularly with an aqueous suspension of hydrocortisone acetate at a rate of 1.42 mg/kg body weight for a period of two weeks. Fibre diameter was measured from clipped tattooed patch wool samples. Follicle activity was measured by histological changes in skin biopsies taken weekly. Blood samples were collected at two-week intervals and plasma cortisol measured. Increased plasma cortisol concentration significantly (p<0.05) reduced clean wool production and mean fibre diameter dropped to its lowest level four weeks after commencement and two weeks after the cessation of cortisol injection. Elevation of plasma cortisol concentration significantly (p<0.0001) increased the percentage of inactive follicles two weeks after injection started. High fibre diameter groups (Strongwool sheep; i.e. HL+HH) had significantly (p<0.0001) higher percentage of follicle shutdown than low fibre diameter groups (Finewool sheep; i.e. LL+LH). Average percentage of shutdown follicles for Finewool (LL+LH) and Strongwool (HL+HH) Merino sheep was $9.8{\pm}0.9$ and $13.5{\pm}0.9$ respectively. Shutdown of primary follicles was more pronounced in Finewool than Strongwool sheep. There was no significant effect of coefficient of variation of fibre diameter on propensity to follicle shutdown induced by exogenous cortisol. It is concluded that elevation in plasma cortisol concentration is inhibitory to the normal activity of follicles in Strongwool sheep but that variation in fibre diameter has little or no effect.

• Biotechnology and Bioprocess Engineering:BBE
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• v.8 no.2
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• pp.151-157
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• 2003

Obtaining a sufficient amount of healthy keratinocytes from a small tissue is difficult. However, ORS cells can be a good source of epithelium since they are easily obtainable and patients do not have to suffer from scar formation at donor sites. Accordingly, the current study modified the conventional primary culture technique to overcome the low propagation and easy aging of epithelial cells during culturing. In a conventional primary culture, the average yield of human ORS tells is 2.↑ $\times$ 10$^3$cells/follicle based on direct incubation in a trypsin (0.1%)/EDTA(0.02%) solution for 15 min at 37$^{\circ}C$, however, our modified method was able to obtain about 6.9 $\times$ 10$^3$cel1s/follicle using a two-step enzyme digestion method involving dispase (1.2 U/mL) and a trypsin (0.1%)/EDTA (0.02%) solution. Thus, the yield of primary cultured ORS cells could be increasd three times higher. Furthermore, a total of 2.0 $\times$ 10$^{7}$ cells was obtained in a serum-free medium. while a modified E-medium with mitomycin C-treated feeder tells produced a total of 6.3 $\times$ 10$^{7}$ Cel1s over 17 days When Starting With 7.5 $\times$ 10$^4$cells. Finally, We Confirmed the effectiveness of our ORS tell isolation method by presenting their ability for reconstructing the bioartificial skin epithelium in vitro

• Korean Journal of Clinical Laboratory Science
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• v.48 no.1
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• pp.8-14
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• 2016

Granulosa cells surround the oocyte within the ovarian follicle and play an essential role in creating conditions required for oocyte as well as follicular development. The current study was conducted to examine the gene expression profile of mouse ovaries during the primordial to primary follicle transition process. Total RNAs from mouse ovaries on day 5 and day 12 were synthesized cDNA using annealing control primers. The DEGs were cloned and their identities were analyzed by BLAST search. The Plekha5 and Anxa11 were highly expressed in primary follicle stage. By contrast, their expression was increased in granulosa cells at the primary follicle stage. We have successfully discovered a list of genes expressed in day 5 and day 12 ovaries and confirmed that some of them are differentially expressed in PMF and/or PRI. This is a spatial-temporal regulatory mechanism on the ovarian folliculogenesis through membrane fusion. The gene expression profile from the current study would provide insight for future study on the mechanism(s) involved in primordial-primary follicular transition. This will provide information for identification of the mechanism of ovarian dysfunction.

• Journal of Embryo Transfer
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• v.26 no.4
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• pp.223-227
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• 2011

The objective of this study was to compare of different isolation method of mouse preantral follicles, and to examine in vitro development of mouse preantral follicles isolated by different method. Preantral follicles were mechanically or enzymatically extracted from mouse ovaries. Mechanical isolation method used fine gauge needles and enzymatic method of isolating follicles used collagenase. The recovered preantral follicles were cultured for 10 days in alpha-minimal essential medium (${\alpha}$-MEM) + 5% FBS + Insulin-Transferrin-Selenium (ITS) + 100 mIU/ml FSH. The collected primary follicles by enzymatic treatment were higher than mechanical method. Others stage preantral follicle by mechanical isolation were higher than enzymatic method. After 10 days of culture, no statistical differences were shown in survival rates of preantral follicle among the 2 culture groups. The metaphase II rates of the oocytes were significantly higher (p<0.05) in mechanical method (17.8%) than in enzymatic method (5.1%). These results suggest that the isolation method of choice depends on the target stage preantral follicles and mechanical isolation is an optimal method of preantral folliclesin a culture of mouse preantral follicle.