• Journal of Periodontal and Implant Science
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• 제52권3호
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• pp.242-257
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• 2022

Purpose: This study investigated periodontal ligament (PDL) restoration in osseointegrated implants using stem cells. Methods: Commercial pure titanium and zirconium oxide (zirconia) were coated with beta-tricalcium phosphate (β-TCP) using a long-pulse Nd:YAG laser (1,064 nm). Isolated bone marrow mesenchymal cells (BMMSCs) from rabbit tibia and femur, isolated PDL stem cells (PDLSCs) from the lower right incisor, and co-cultured BMMSCs and PDLSCs were tested for periostin markers using an immunofluorescent assay. Implants with 3D-engineered tissue were implanted into the lower right central incisors after extraction from rabbits. Forty implants (Ti or zirconia) were subdivided according to the duration of implantation (healing period: 45 or 90 days). Each subgroup (20 implants) was subdivided into 4 groups (without cells, PDLSC sheets, BMMSC sheets, and co-culture cell sheets). All groups underwent histological testing involving haematoxylin and eosin staining and immunohistochemistry, stereoscopic analysis to measure the PDL width, and field emission scanning electron microscopy (FESEM). The natural lower central incisors were used as controls. Results: The BMMSCs co-cultured with PDLSCs generated a well-formed PDL tissue that exhibited positive periostin expression. Histological analysis showed that the implantation of coated (Ti and zirconia) dental implants without a cell sheet resulted in a well-osseointegrated implant at both healing intervals, which was confirmed with FESEM analysis and negative periostin expression. The mesenchymal tissue structured from PDLSCs only or co-cultured (BMMSCs and PDLSCs) could form a natural periodontal tissue with no significant difference between Ti and zirconia implants, consequently forming a biohybrid dental implant. Green fluorescence for periostin was clearly detected around the biohybrid implants after 45 and 90 days. FESEM showed the invasion of PDL-like fibres perpendicular to the cementum of the bio-hybrid implants. Conclusions: β-TCP-coated (Ti and zirconia) implants generated periodontal tissue and formed biohybrid implants when mesenchymal-tissue-layered cell sheets were isolated from PDLSCs alone or co-cultured BMMSCs and PDLSCs.

• BMB Reports
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• 제56권3호
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• pp.202-207
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• 2023

We investigated the neuroprotective effects of deca nano-graphene oxide (daNGO) against reactive oxygen species (ROS) and inflammation in the human neuroblastoma cell line SH-SY5Y and in the 6-hydroxydopamine (6-OHDA) induced Parkinsonian rat model. An MTT assay was performed to measure cell viability in vitro in the presence of 6-OHDA and/or daNGO. The intracellular ROS level was quantified using 2',7'-dichlorofluorescein diacetate. daNGO showed neuroprotective effects against 6-OHDA-induced toxicity and also displayed ROS scavenging properties. We then tested the protective effects of daNGO against 6-OHDA induced toxicity in a rat model. Stepping tests showed that the akinesia symptoms were improved in the daNGO group compared to the control group. Moreover, in an apomorphine-induced rotation test, the number of net contralateral rotations was decreased in the daNGO group compared to the control group. By immunofluorescent staining, the animals in the daNGO group had more tyrosine hydroxylase-positive cells than the controls. By anti-Iba1 staining, 6-OHDA induced microglial activation showed a significantly decrease in the daNGO group, indicating that the neuroprotective effects of graphene resulted from anti-inflammation. In conclusion, nano-graphene oxide has neuroprotective effects against the neurotoxin induced by 6-OHDA on dopaminergic neurons.

• Pediatric Infection and Vaccine
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• 제9권1호
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• pp.67-73
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• 2002

목 적 : 최근 2~3년 주기로 홍역이 유행하고 있으며, 특히 2000년 말부터 2001년 초까지는 홍역의 집단발생으로 방역당국과 의료계가 비상이었다. 이에 저자들은 홍역에 이환된 환아의 혈청 항홍역 항체를 측정하고 홍역 예방 접종 유무에 따른 차이를 조사하여, 홍역 예방 접종의 효과를 보고, 추가접종의 필요성 및 시기를 살펴보고자 본 연구를 시행하였다. 방 법 : 2000년 6월부터 2001년 6월까지 임상적으로 홍역이 의심되어 원주기독병원에 입원 또는 외래 방문한 298명의 환아 중, criteria for disease control을 만족하는 201명의 환아를 대상으로 홍역 발생의 성별, 월별분포, 예방접종 유무에 따른 연령별 분포 및 항홍역 항체의 양상을 조사하였다. 항홍역 항체의 측정은 발진시기에 채혈한 혈액을 Immunofluorescent Assay로 시행하였으며 IgG 항체는 역가가 1 : 10 이상인 경우를 양성으로 판정하였고, IgM 항체는 양성인 경우를 양성으로 하였다. 결 과 : 1) 대상군은 남아 117명, 여아 84명으로 남녀비는 1.4 : 1이었다. 2) 월병 발생 빈도는 5월에 38례(18.9%)로 가장 많이 발생하였으며, 2000년 11월부터 2001년 1월까지 증가 추세에 있다가 2월과 3월에 감소한 후 4월부터 다시 증가하였다. 3) 대상군 중 예방 접종을 시행한 경우는 93례(46.3%), 시행하지 않은 경우는 108례(53.7%)로 비접종군이 더 많았으나, 통계학적으로 유의한 차이는 없었다. 4) 접종군에서의 연령별 분포는 10세 이상에서 54례(58.1%), 7~10세가 15례(16.0%), 15개월~3세가 12례(12.9%), 4~6세 및 6~14개월이 각각 6례(6.5%)였다. 5) 비접종군에서의 연령별 분포는 6~14개월이 88례(81.5%), 15개월~3세가 9례(8.3%), 6개월 미만이 7례(6.5%), 10세 이상 3례(2.8%), 7~10세가 1례(0.9%)였다. 6) 예방접종 유무에 따른 항홍역 항체 IgG, IgM의 분포는 접종군에서 IgG, IgM 모두 양성인 경우가 78례(87.6%), IgG(+) IgM(-)인 경우가 11례(12.4%)였으며, 비접종군에서는 IgG, IgM 모두 양성인 경우가 69례(63.9%), IgG(-), IgM(+)인 경우는 39례(36.1%)로 나타났다. 결 론 : 홍역에 이환된 환아 중 비접종군에서는 6~14개월에 이환율이 가장 높았으며 이는 6~14개월에 모체로부터 받은 항체가 가장 많이 소실되기 때문으로 사료된다. 접종군에서는 10세 이상에서 이환율이 가장 높았고, IgG, IgM 모두 양성이면서 홍역에 이환된 경우가 대부분인 점으로 보아, 홍역 특이 IgG 항체 역가가 10세 이후에 가장 많이 감소하는 것으로 추측할 수 있다. 따라서 홍역의 퇴치를 위해서 홍역의 초기 예방접종 뿐만 아니라 재접종이 중요하며, 특히 재접종의 시기가 매우 중요할 것으로 사료된다.

• Tuberculosis and Respiratory Diseases
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• 제54권5호
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• pp.495-509
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• 2003

연구배경 : 호중구에 의해 매개되는 염증반응은 호중구의 수명이 매우 짧기 때문에 대부분 자연 종결된다. 패혈증에서는 이러한 호중구의 아포프토시스가 감소되어 수명이 연장되어 있어서 지속적인 염증반응이 일어나게 된다. 호중구의 수명 연장을 유도하는 많은 염증 매개 물질들이 nuclear factor-${\kappa}B$ 전사인자에 의해 조절되기 때문에 이 연구에서는 nuclear factor-${\kappa}B$가 패혈증 환자에서 관찰되는 호중 구의 아포프토시스 억제와 연관이 있을 것으로 가정하였다. 방법 : 건강한 정상인과 패혈증 환자의 호중구를 정맥혈로부터 신선하게 분리하여 실험하였다. 호중구의 아포프토시스는 특징적인 아포프토시스의 형태를 보이는 세포를 광학현미경으로 세거나 Annexin V를 이용한 유세포분석법으로 정량하였다. Nuclear factor-${\kappa}B$의 활성도는 면역형광 엽색법 또는 electrophoretic mobility shift assay로 판단하였다. 항아포프토시스 단백인 X-linked inhibitor of apoptosis의 발현 정도는 western blot으로 평가하였다. 결과 : 패혈증 환자에서 자발적 아포프토시스가 감소되어 있음을 확인하였다. 패혈증 환자의 호중구에 cycloheximide를 처치하였을 때 아포프토시스가 유의하게 증가하여 아포프토시스 감소가 새로운 단백 합성에 의존적임을 관찰하였다. 패혈증 환자의 호중구에서는 정상인과는 달리 기저상태에서도 nuclear factor-${\kappa}B$가 핵 내로 이동되어 활성화되어 있었고 nuclear factor-${\kappa}B$의 활성을 억제하였을 때 아포프토시스의 억제가 반전되었다. 또한 nuclear factor-${\kappa}B$에 의존적인 X-linked inhibitor of apoptosis 단백의 발현 수준이 정상인의 호중구에서는 24시간 동안 배양하면서 점차 감소하였지만 패혈증 환자의 호중구에서는 지속적으로 발현 수준이 유지되었다. 결론 : 패혈증 환자에서 관찰되는 호중구의 아포프토시스 억제에는 nuclear factor-${\kappa}B$ 전사인자의 활성화에 의한 생존 단백의 유도가 관여할 것으로 생각하였다.

• Maxillofacial Plastic and Reconstructive Surgery
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• 제32권2호
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• pp.97-106
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• 2010

Aim of the study: An alternative source of adult stem cells that could be obtained in large quantities, under local anesthesia, with minimal discomfort would be advantageous. Adipose tissue could be processed to obtain a fibroblast-like population of cells or adipose tissue-derived stromal cells (ATSCs). This study was performed to confirm the availability of ATSCs in bone tissue engineering. Materials amp; Methods: In this study, adipose tissue-derived mesenchymal stem cell was extracted from the liposuctioned abdominal fat of 24-old human and cultivated, and the stem cell surface markers of CD 105 and SCF-R were confirmed by immunofluorescent staining. The proliferation of bone marrow mesenchymal stem cell and ATSCs were compared, and evaluated the osteogenic differentiation of ATSCs in a specific osteogenic induction medium. Osteogenic differentiation was assessed by von Kossa and alkaline phosphatase staining. Expression of osteocyte specific BMP-2, ALP, Cbfa-1, Osteopontin and osteocalcin were confirmed by RT-PCR. With differentiation of ATSCs, calcium concentration was assayed, and osteocalcin was evaluated by ELISA (Enzyme-linked immunosorbant assay). The bone formation by 5-week implantation of HA/TCP block loaded with bone marrow mesenchymal stem cells and ATSCs in the subcutaneous pocket of nude mouse was evaluated by histologic analysis. Results: ATSCs incubated in the osteogenic medium were stained positively for von Kossa and alkaline phosphatase staining. Expression of osteocyte specific genes was also detected. ATSCs could be easily identified through fluorescence microscopy, and bone formation in vivo was confirmed by using ATSC-loaded HA/TCP scaffold. Conclusions: The present results show that ATSCs have an ability to differentiate into osteoblasts and formed bone in vitro and in vivo. So ATSCs may be an ideal source for further experiments on stem cell biology and bone tissue engineering.

• Pediatric Infection and Vaccine
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• 제12권2호
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• pp.108-113
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• 2005

목 적 : 최근 HHV-8는 면역력이 정상인 소아에서 원발성 감염에 의해 발진이 동반된 발열성 질환을 유발하는 것으로 알려지고 있다. 본 연구는 국내 정상 소아의 항 HHV-8 항체 양성률을 알아보기 위해 시행되었다. 방 법 : 항 HHV-8 항체의 양성률을 파악하기 위하여 인제의대 상계백병원을 방문한 환자 중에서, 최근 3주간 발열 증상이 없었던 건강한 소아 112명의 혈청을 대상으로 하였다. Lytic 바이러스 항원에 대한 IgG 항체 검사는 간접면역 형광검사법, HHV-8의 ORF에 대한 특이 항체 검사는 peptide mix ELISA 검사법을 사용하였다. 결 과 : 대상 환아는 총 112명이었으며 남아 64명, 여아 48명이었다. 1세 이하의 연령군은 17명이었으며, 2~5세 연령군 24명, 6~9세 연령군 24명, 10~15세 연령군은 47명이었다. HHV-8에 대한 혈청 항체 양성률은 간접 면역 형광검사법으로 3.5%(4/112), ELSIA로는 1.8%(2/112)였다. 결 론 : 국내 소아의 HHV-8 항체의 양성률은 비교적 낮았지만 앞으로 발진이 동반된 발열성 질환에서 HHV-8의 역할에 대한 추가적인 연구가 필요할 것으로 보인다.

• 한국가축번식학회지
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• 제13권2호
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• pp.113-120
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• 1989

In order to determine the sex of preimplantation embryos prior to transfer in cattle, a series of experiments were carried out using 45 Holstein donor cows to examine the ovarian response on the gonadotropin and PGF2${\alpha}$, and the morphology of fresh embryos or frozen/thawed embryos after deep freezing at -196$^{\circ}C$. The sexing of embryos treated with the medium containing H-Y antiserum(10%, v/v) and FITC anti-mouse IgG(10%, v/v) were analysed by chromosomal analysis, and the sex of the embryos which survived were ascertain after delivering the pups. The results obtained were summarized as follows ; 1. The average number of developed follicle and corpus luteum per cow were 13.5 and 8.1, and the ovalation rate was 60.1%. 2. Of 220-ova recovered, 75(34.1%) were morula and 91(41.4%) were blastocyst, and the morphological normal and abnormal rate of ova recovered were 75.5% and 24.5%, respectively. 3. Of 39 frozen/thawed embryos, the scores of normal morula and blastocyst, after thawing were 79.2%(19/24) and 73.3%(11/15). The average rate of frozen/thawed embryos which appeared morphologically normal post thawing was 76.9%(30/39). 4. The sex ratio was measured using the embryos treated with immunofluorescence assay to examine the relationship between embryo developmental stage, sex ratio of morula stage embryo was 42.2%(19/45) fluorescing and 57.8%(26/45) non-fluorescing, on the other hand, the ratio switched to 46.8%(29/62) fluorescing and 53.2%(33/62) non-fluorescing embryo in blastocyst stage. The sex ratio was also measured between fresh and frozen/thawed embryos, fresh and frozen/thawed treated embryos were indicated 45.8%(38/83) fluorescing, 54.2%(45/83) non-fluorescing and 41.7%(10/24) fluorescing, 58.3%(14/24) non-fluorescing. This trend indicated the approximal sex ratio was 1 : 1. 5. The result of karyotype test showed the successful rate of sexing embryo is fluorescing and non-fluorescing was 21.2%(7/33) and 29.6%(8/27). The female to male ratio within 33 fluorescing was 28.6 : 71.4, and the ratio of 27 non-fluorescing embryos was 87.7 : 12.5. 6. Of the embryo transferred after assignment of H-Y phenotype, five of the fluorescing embryos survived to term, all was males. Whereas six non-fluorescing embryos also survived to term and the sexes of the calves were 1 male 5 female.

• Reproductive and Developmental Biology
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• 제36권1호
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• pp.33-37
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• 2012

Differential capacity of the parthenogenetic embryonic stem cells (PESCs) is still under controversy and the mechanisms of its neural induction are yet poorly understood. Here we demonstrated neural lineage induction of PESCs by addition of insulin-like growth factor-2 (Igf2), which is an important factor for embryo organ development and a paternally expressed imprinting gene. Murine PESCs were aggregated to embryoid bodies (EBs) by suspension culture under the leukemia inhibitory factor-free condition for 4 days. To test the effect of exogenous Igf2, 30 ng/ml of Igf2 was supplemented to EBs induction medium. Then neural induction was carried out with serum-free medium containing insulin, transferrin, selenium, and fibronectin complex (ITSFn) for 12 days. Normal murine embryonic stem cells derived from fertilized embryos (ESCs) were used as the control group. Neural potential of differentiated PESCs and ESCs were analyzed by immunofluorescent labeling and real-time PCR assay (Nestin, neural progenitor marker; Tuj1, neuronal cell marker; GFAP, glial cell marker). The differentiated cells from both ESC and PESC showed heterogeneous population of Nestin, Tuj1, and GFAP positive cells. In terms of the level of gene expression, PESC showed 4 times higher level of GFAP expression than ESCs. After exposure to Igf2, the expression level of GFAP decreased both in derivatives of PESCs and ESCs. Interestingly, the expression level of $Tuj1$ increased only in ESCs, not in PESCs. The results show that IGF2 is a positive effector for suppressing over-expressed glial differentiation during neural induction of PESCs and for promoting neuronal differentiation of ESCs, while exogenous Igf2 could not accelerate the neuronal differentiation of PESCs. Although exogenous Igf2 promotes neuronal differentiation of normal ESCs, expression of endogenous $Igf2$ may be critical for initiating neuronal differentiation of pluripotent stem cells. The findings may contribute to understanding of the relationship between imprinting mechanism and neural differentiation and its application to neural tissue repair in the future.

• 한국동물학회지
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• 제31권1호
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• pp.49-55
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• 1988

SV80와 같은 SV40로 형질전환된 사람세포는 종양을 일으킬 수 있는 능력을 가지고 있으나 면역기구인 흉선이 없는 누드마우스에서는 거부반응을 일으켜 종양을 일으키지 않는다. 그러나, 예외적으로 WI18/VA-2세포는 누드마우스에서 종양을 일으키며 이에서 얻는 두클론중 NW18C11은 종양을 일으키나 NW18C12는 종양을 일으키지 않는다. 본 실험에서는 이들 두 클론의 차이점들을 조사하였다. 실험결과, NW18C11은 NW18C12보다 더 많은수의 SV40 sequence를 포함하고 있음을 southern blot방법을 통해 확인하였으며 또한 immunofluoresce와 immunoprecipitation방법을 사용하여 두 클론 모두 정상크기의 SV40유전자산물인 large T와 small t 단백질을 생성함을 확인하였다. 한편 두 클론내에 포함되어 있는 바이러스유전자가 비형질전환새포로 하여금 생체내에서 악성종양 형성능력을 획득하도록 형질전환시킬수 있는지 확인하기 위해 두 클론의 DNA를 추출하여 마우스 NIH3T3세포에 주입시켜 형질전환된 세포를 선별하였다. 이 세포들은 모두 large T단백질을 생성하였으며 누드마우스에서 종양을 일으켰다. 이들 결과로써 NW18C12세포의 형질전환능은 완전하며, 이 세포가 누드마우스에서 거부반응에 기인하는 것으로 생각된다.

• Journal of Microbiology and Biotechnology
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• 제18권7호
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• pp.1308-1316
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• 2008

Propionibacterium acnes is known to playa pivotal role in the pathogenesis of acne vulgaris. CBT-SL5 is one of the antimicrobial peptides from Enterococcus faecalis SL5, and it has shown antimicrobial activity against P. acnes. The aim of this study was to investigate the anti-inflammatory effect of CBT-SL5 on the inflammation induced by P. acnes in cultured human keratinocyes. Cultured human keratinocytes derived from neonatal foreskin were treated with heat-killed P. acnes to induce inflammation, and then various concentrations of CBT-SL5 were added to the P. acnes-treated keratinocytes. The mRNA expression and protein secretion of interleukin (IL)-8, an inflammation marker, was analyzed by real-time reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. We also analyzed the nuclear factor-kappa B (NF-$\kappaB$) p65 translocation by performing immunofluorescent staining. P. acnes treatment up regulated the IL-8 mRNA expression in the keratinocytes, and this was brought about through both toll-like receptor (TLR)2 and TLR4. At the concentrations of 10, 50, and 100 ng/ml, CBT-SL5 significantly down regulated the P. acnes-induced IL-8 mRNA expression and protein production (p<0.05). At 6 hand 12 h of the treatment, CBT-SL5 significantly suppressed the P. acnes-induced IL-8 mRNA expression. Secretion of IL-8 protein was significantly reduced at 24 h. The functional inhibitory activity of CBT-SL5 was shown by CBT-SL5 suppressing the P. acnes-induced NF-$\kappaB$ translocation from the cytoplasm to the nucleus. These results demonstrated that CBT-SL5 suppressed the P. acnes-induced IL-8 expression in keratinocytes. Therefore, CBT-SL5 may be a novel anti-inflammatory treatment for acne.