凝集素在面颈部皮肤色素痣和恶性黑色素瘤中的细胞定位

观察了10种凝集素在面颈部皮肤色素痣(37例)和恶性黑色素瘤(10例)中细胞定位。除WGA标记痣细胞外,其余9种凝集素染色均为阴性。LCA、PNA、PSA、PHA、RCA_1和WGA标记部分恶黑细胞,但凝集素受体的分布不同于痣细胞。结果表明痣细胞和恶黑之间,其细胞表面糖基表达有很大差别。     

CM-DiI体外标记人乳牙牙髓干细胞的可行性研究

目的: 筛选CM-DiI用于人乳牙牙髓干细胞标记的最佳浓度,并对标记后细胞的生物学行为进行评价,为下一步细胞活体移植和体内示踪打下基础。方法: 酶解组织块法培养人乳牙牙髓细胞并纯化,免疫组化鉴定细胞来源,进行细胞体外多向诱导分化能力实验。将第3代细胞分别使用浓度2 mg/L、3 mg/L及4 mg/L的CM-DiI进行标记,比较标记后细胞乳酸脱氢酶释放率及细胞增殖情况,并观察经体外多次传代后细胞荧光的表达情况。结果: 酶解组织块法可获得成集落状生长的人乳牙牙髓细胞;免疫组化确定细胞为间充质而非造血来源;细胞具有成脂、成骨及成牙本质等多向分化能力。3种标记物浓度均未对细胞乳酸脱氢酶释放及细胞增殖产生显著的不利影响;随体外多次传代,标记荧光强度逐渐减退。结论: CM-DiI操作简便、染色速度快,可有效的标记人乳牙牙髓干细胞,其中3 mg/L的标记浓度细胞毒性极小,标记效率高,在体外多次传代后荧光信号仍能稳定表达。     

颌骨成釉细胞瘤中细胞凋亡相关蛋白的表达

目的研究颌骨成釉细胞瘤中细胞凋亡相关蛋白的表达。方法采用免疫组化方法观察39例经典型成釉细胞瘤中凋亡相关蛋白Fas、Fas配体（Fas Ligand，FasL）、caspase-3和bcl-2及增殖细胞标记Ki67的表达，采用TUNEL法检测11例成釉细胞瘤中的凋亡细胞。结果39例成釉细胞瘤中分别有35（89．7％）、34（87．2％）及21（61．5％）例表达Fas、FasL和caspase-3，阳性染色主要分布于上皮岛中心呈鳞状化生和颗粒变性的细胞，TUNEL法标记的凋亡细胞也集中于这些区域。bcl-2和Ki67阳性染色则主要位于上皮岛周边的基底细胞。结论Fas／FasL介导的细胞凋亡机制可能在成釉细胞瘤中肿瘤细胞的分化、变性及被清除过程中发挥作用。     

功能矫形前伸下颌后对大鼠髁突软骨DNA合成影响的研究

应用放射性自显影技术,以~3H-TdR 为标记物,观察了下颌功能性前仲后大鼠髁突软骨细胞DNA 合成的适应性变化。结果发现下颌前伸后,改变了 TMJ 的生物力学和生物物理环境,刺激了 G_0期细胞进入细胞增殖周期,因而髁突软骨生发层内~3H-TdR 标记细胞增多。成熟层~3H-TdR 标记细胞增多,是由于生发层细胞有丝分裂增加,大量标记的 G_1期细胞进入了分化阶段,继发性地使成熟层内标记细胞增多。     

涎腺原发恶性肌上皮瘤19例的病理诊断分析

目的 探讨涎腺多结节形态原发恶性肌上皮瘤的病理特征。方法 光镜观察19例涎腺多结节形态原发恶性肌上皮瘤的组织学、细胞学结构,其中11例做Calponin、平滑肌肌动蛋白(SMA)、S-100、神经胶质酸性蛋白(GFAP)、细胞角蛋白、增殖细胞核抗原(PCNA)的免疫组化检测,3例做超微结构观察。结果 肿瘤上皮巢呈多结节生长,中央常伴坏死,肿瘤侵袭周围组织,细胞有异形,肿瘤细胞主要为上皮样细胞型,有较多肿瘤性基质形成。肌上皮性标记及部分细胞角蛋白、PCNA标记阳性。电镜下见肿瘤细胞内有肌微丝。结论 免疫组化染色及超微结构观察均支持肿瘤为肌上皮来源及恶性特征。     

CM-Dil标记人牙龈间充质干细胞在大鼠颅骨缺损修复模型中的示踪研究

目的:探讨氯甲基苯甲酰氨(CM-Dil)荧光染料体外标记人牙龈间充质干细胞(hGMSCs)对其增殖、分化能力的影响以及在体内荧光标记持续时间.方法:组织块法原代培养hGMSCs,采用流式细胞术检测间充质干细胞表面标志物,然后以CM-Dil标记hGMSCs,CCK-8检测标记后细胞的增殖;标记及未标记的细胞进行成骨及成脂...     

Smad 2/3、7蛋白在OLP CD8+T细胞中的表达

目的:观察Smad2/3、在口腔扁平苔藓(oral lichen planus,OLP)CD8 T细胞中的表达。方法:采用免疫7 组化双标记法检测28例OLP组织中CD8 T细胞Smad2/3、7蛋白表达水平。设10例临床正常口腔黏膜(Normal oral mucosa,)为对照组。结果:NOMNOM组织中CD8 /Smad2/3、CD8 /Smad7双标记阳性细胞均为零。OLP组织中, CD8/Smad2、3和CD8 /Smad7双标记细胞的阳性率分别为1.020 1.24,5.524±5.074。结论:与NOM相比, OLP组织中CD8 T细胞Smad2、 3表达无明显升高。而Smad7阳性颗粒均位于细胞浆中,可能部分阻碍了TGF-β/Smad正向反馈通路,从而引起CD8 T细胞抑制不足,造成OLP慢性炎症的长期持续。     

转化生长因子β受体在口腔扁平苔藓CD8+T细胞中的表达

目的 研究转化生长因子β受体(transforming growth factor-β receptor,TGF-βR)在口腔扁平苔藓(oral lichen planus,OLP)组织中CD8+T细胞中的表达,探讨TGF-βR在OLP发病中的作用.方法 采用免疫组化双标记法检测28例OLP组织中CD8+T细胞TGF-βR Ⅰ和TGF-βR Ⅱ蛋白的表达.以10例临床正常口腔黏膜(normal oral mucosa,NOM)为对照组.结果 OLP组织中CD8+及TGF-βR Ⅰ、CD8+及TGF-βR Ⅱ双标记细胞的阳性率分别为(8.82±9.98)%、(1.11±2.94)%.NOM组织中双标记细胞均为零.OLP组CD8+T细胞中TGF-βRⅡ表达与NOM组相比差异无统计学意义(P＞0.05),TGF-βR Ⅰ与NOM组相比表达增强.结论 提示OLP组织CD8+T细胞存在TGF-β信号传导异常.这可能是CD8+T细胞缺乏有效抑制、致使OLP慢性炎性反应长期持续的因素之一.     

金地鼠颊囊癌前病变的细胞动力学研究

采用放射性自显影技术~3H—Tdr两次标记法研究了DMBA诱发的金地鼠颊囊癌前病变的细胞动力学改变。测定结果发现,随癌前病变的发展,细胞增殖周期缩短、标记指数增高。标记细胞在正常上皮仅出现于基底层,于临近癌变前扩散进入棘层。结果证实随癌前病变的发展、细胞增长速度加快,细胞分化成熟障碍。     

重组酶Cre在体外对LoxP标记的基因组进行重组的初步研究

目的：研究逆转录病毒体系转导的重组酶Cre在体外对LoxP标记的基因组进行重组的能力。方法：用逆转录病毒表达Cre,体外感染条件性基因剔除（即LoxP标记的基因）小鼠来源的原代培养的牙乳头间充质细胞。从被感染的牙乳头间充质细胞中提取基因组DNA,用PCR的方法显示LoxP标记的基因组长度的变化,从而反映Cre在体外的重组能力。结果：被表达Cre的逆转录病毒感染的牙乳头间充质细胞,其标记的基因组长度发生了预期的变化,被标记的部分发生缺失。结论：用逆转录病毒体系转导重组酶Cre吸效地对LoxP标记的基因组进行重组。为用这一方法在体外研究特异基因对细胞生物学行为的影响奠定基础。     

Extent and diversity of inflammatory cell infiltrates in squamous cell carcinomas and basal cell epitheliomas of the head and neck

Using monoclonal antibodies reactive with Langerhans' cells (LCs), macrophages, and T cell subpopulations, the density and proportions of cells of the immune system of the normal oral mucosa were determined immunohistochemically, and compared with findings in oral squamous cell carcinomas (SCC) and basal cell epitheliomas (BCE). In normal oral epithelia, the dominant cell type was the LC, positive for CD 1, and expressing HLA-DR antigens (DR+). Many intraepithelial cells were lymphocytes of the suppressor/cytotoxic phenotype (CD 8+), which was also the most prominent cell type in the normal mucosal stroma. Significant differences were observed for the content of CD 8-, OKM 1-, and CD 4-positive cells in the epithelium of normal oral mucosa, SCC, and BCE, and for the amount of CD 1-positive Langerhans cells in the connective tissue of the different groups of tissues. When CD 4/CD 8 ratios were calculated, differences between SCC and BCE became most evident. A CD 4/CD 8 ratio greater 0.5 was seen to be characteristic for BCE. Thus, in contrast to the striking preponderance of suppressor/cytotoxic lymphocytes (CD 8+) in SCC, BCE showed typically almost balanced numbers of suppressor/cytotoxic (CD 8+) and helper/inducer (CD 4+) lymphocytes. This finding further underlines the biological differences recognized between these most common neoplasias of the head and neck.     

Extent and diversity of inflammatory cell infiltrates in squamous cell carcinomas and basal cell epitheliomas of the head and neck

Using monoclonal antibodies reactive with Langerhans' cells (LCs), macrophages, and T cell subpopulations, the density and proportions of cells of the immune system of the normal oral mucosa were determined immunohistochemically, and compared with findings in oral squamous cell carcinomas (SCC) and basal cell epitheliomas (BCE). In normal oral epithelia, the dominant cell type was the LC, positive for CD 1, and expressing HLA-DR antigens (DR⁺). Many intraepithelial cells were lymphocytes of the suppressor/cytotoxic phenotype (CD 8⁺), which was also the most prominent cell type in the normal mueosal slroma. Significant differences were observed for the content of CD 8-, OKM 1-, and CD 4-positive cells in the epithelium of normal oral mucosa, SCC, and BCE, and for the amount of CD 1-positive Langerhans cells in the connective tissue of the different groups of tissues. When CD 4/CD 8 ratios were calculated, differences between SCC and BCE became most evident. A CD 4/CD 8 ratio greater 0.5 was seen to be characteristic for BCE. Thus, in contrast to the striking preponderance of suppressor/cytotoxic lymphocytes (CD 8⁺) in SCC, BCE showed typically almost balanced numbers of suppressor/cytotoxic (CD 8⁺) and helper/induced (CD 4⁺) lymphocytes. This finding further underlines the biological differences recognized between these most common neoplasias of the head and neck.     

An immunohistochemical study of oral submucous fibrosis

Oral submucous fibrosis (OSF) is a chronic disease of the oral cavity characterized by inflammation and progressive mucosal fibrosis. These reactions may be the result of either direct stimulation from exogenous antigens like areca alkaloids or by changes in tissue antigenicity that may lead to an autoimmune response. This study investigated the presence and distribution of inflammatory cells and MHC class II antigen expression by epithelial and immunocompetent cells using a three-stage immunoperoxidase method on frozen sections. Thirty OSF tissue specimens and ten normal buccal mucosae were studied and compared. All tissues were investigated using antibodies to T cells (CD3), T helper/inducer cells (CD4), T suppressor/cytotoxic cells (CD8), B cells (CD20), naive T cells and monocytes (CD45RA), macrophages. Langerhans' cells (CD68) and HLA-DR-positive cells (HLA-DR alpha). The predominant cell populations detected in normal tissues were CD3, CD4 and HLA-DR-positive cells. The distribution of CD4-positive cells was similar to that of CD3-positive cells, which were scattered, often uniformly distributed, both in the epithelium and connective tissue. CD8-positive cells were occasionally seen in the normal epithelium and lamina propria. Few scattered B cells (CD20) and macrophages (CD681) were observed in normal mucosa. Naive T cells (CD45RA) were seen in all normal tissues focally concentrated around the connective tissue papillae with a similar distribution to that of CD3-positive cells. All normal sections showed HLA-DR-positive cells scattered both in the epithelium and in the lamina propria. Epithelial cells did not show any positive reaction to this antibody and many intraepithelial positive cells showed a dendritic morphology. The cell populations detected in OSF showed higher numbers of CD3 and HLA-DR-positive cells compared with those of the normal tissues. The pattern of staining for CD4-positive cells in OSF tissues was similar to that of CD3-positive cells both in the epithelium and connective tissue and was higher than that in normal tissues. A few scattered CD8-positive cells and only occasional CD20- and CD68-positive cells were seen in OSF sections. Few CD45RA-positive cells were found in the epithelium and lamina propria of OSF sections. However, OSF specimens showed high numbers of HLA-DR-positive cells in the basal layer of the epithelium, juxtaepithelium and in the lamina propria in a similar distribution to that of CD3 cells compared with the normal tissues. Most HLA-DR-positive cells in the epithelium showed dendrites directed vertically towards the surface. The increased evidence of CD4 and HLA-DR-positive cells in OSF tissues suggests that most lymphocytes were activated and shows an increased presence of Langerhans' cells. The presence of these immunocompetent cells and high ratio of CD4 to CD8 in OSF tissues suggest an ongoing cellular immune response leading to a possible imbalance of immunoregulation and alteration in local tissue architecture.     

Lamina propria dendritic cells express activation markers and contact lymphocytes in chronic periodontitis

BACKGROUND: Dendritic cells are characterized by shape, structure, and membrane molecule expression; they contact T lymphocytes to present antigens and stimulate plasma cell differentiation in vitro. Dendritic cells are known to be present in healthy human gingiva and to be altered in HIV-associated periodontitis. Here, we address the phenotype, location, and intercellular relationships of dendritic cells in chronic periodontitis. METHODS: Biopsies from patients with chronic periodontitis were analyzed by electron microscopy and indirect immunofluorescence for dendritic cells and lymphocyte markers. RESULTS: Langerhans' cells were spread in oral epithelium but restricted to the basal layer in pocket epithelium; they did not usually express major histocompatibility complex (MHC)-II antigens nor contact lymphocytes. Dendritic cells were abundant in the lamina propria of pocket epithelium; they were MHC-II positive, admixed with CD4-positive and CD8-positive T lymphocytes, and, they expressed CD54, CD80, and CD86. Dendritic cells often contacted lymphocytes and were also located within plasma cell aggregates. CONCLUSIONS: The data suggest that prerequisites for mounting a T cell-mediated immune response exist in chronic periodontitis, although this response is limited to the lamina propria. These results suggest that T-cell responses offer limited protection and can contribute to tissue damage during periodontal disease.     

Chemokine expression in Porphyromonas gingivalis-specific T-cell lines

Autologous non-T cells (monocytes and B cells) were added to Porphyromonas gingivalis-specific T cell lines established from 9 healthy adults together with P. gingivalis outer membrane antigens for 4-6, 16-18, 24 and 48 h. Flow cytometry was employed to analyze the CD4 and CD8 cells, monocytes and B cells for intracytoplasmic IP-10 (interferon-gamma inducible protein 10), MCP-1 (monocyte chemoattractant protein 1), MIP-1 alpha (macrophage inflammatory protein 1 alpha) and RANTES (regulated on activation normal T cell expressed and secreted) at the four time periods. All cell types were positive for each chemokine throughout the 48-h time period. There were significantly fewer MCP-1-positive cells compared with the other 3 chemokines. However, the percentages of MCP-1, MIP-1 alpha- and RANTES-positive CD8 cells were significantly higher than the percentages of positive CD4 cells in all cultures. IP-10-positive CD4, CD14-positive monocytes and CD19-positive B cells were predominant compared with MIP-1 alpha- and RANTES-positive cells at 24 h. In conclusion, the present study has shown that P. gingivalis-specific T cells, monocytes and B cells produce chemokines in response to P. gingivalis outer membrane antigens, IP-10 being predominant, with MCP-1 being significantly reduced in comparison with IP-10, MIP-1 alpha and RANTES. Increased percentages of CD8 cells were induced to produce chemokines in comparison with CD4 cells, indicating a more preferential action on CD8 rather than CD4 cells.     

Quantitative analysis of the immunocompetent cells in periapical granuloma: correlation with the histological characteristics of the lesions

Granuloma formation includes an immune response in oral tissues to various microorganisms and their products. The immunocompetent cells of both series (T and B) are present in the periapical lesions. In order to further analyze the relative contribution and pathophysiological significance of the T cell subsets in granuloma formation, we undertook the quantitative analysis of the CD3-positive, CD4-positive, CD8-positive and Ig-positive cells in these lesions by using indirect immunofluorescence. Evidence is provided showing predominance of T cells in diffuse and B cells in focal mononuclear infiltrates. CD8-positive cells were more frequent in diffuse infiltrates and in particular in granulomas with distinct epithelium while CD4-positive cells were more numerous in focal infiltrates. It appears that the presence and ratios of different subsets of immunocompetent cells reflects the pathogenesis of granuloma and transformation to cyst.     

Immunohistochemical analysis of CD1a-labeled Langerhans cells in human dental periapical inflammatory lesions--correlation with inflammatory cells and epithelial cells

OBJECTIVE: Distribution and density of CD1a-labeled Langerhans cells (LCs) were examined in human dental periapical inflammatory lesions, and compared with inflammatory cell infiltration or epithelial cell proliferation. MATERIALS AND METHODS: Eighty three periapical lesions (26 apical granulomas (AGs), 8 epitheliated granulomas (EGs), 34 radicular cysts (RCs), 15 residual radicular cysts (RRCs)) were collected. As control, specimens of periodontal ligaments including Malassez epithelial rests were curetted from 21 teeth. LC densities were measured and various degrees of inflammatory cell infiltration were examined immunohistochemically. Labeling indices for the cellular proliferation markers Ki-67 antigen and DNA topoisomerase II alpha were calculated in the epithelial components. RESULTS: LCs were found in all periapical lesions but not in Malassez epithelial rests. LCs were more abundant in epithelial components than in subepithelial layers. Intraepithelial LCs were more frequent in RCs than in RRCs, whereas subepithelial LCs were less frequent in RRCs than in AGs and EGs. T lymphocytes consistently outnumbered macrophages, plasma cells and B lymphocytes. The range of the CD4/CD8-positive cell ratio differed according to the lesions. Increased LC density was associated with the severity of CD3-positive cell infiltration. Ki-67- and Topo II-LI showed various degrees of epithelial immunoreactivity. There was a significant correlation between LC density and proliferative potential of the epithelium in periapical lesions. CONCLUSION: These findings suggested that LCs appeared to be associated with T lymphocyte infiltration and proliferative potential of the epithelial tissue in periapical lesions.     

The proportion of interleukin-4, interferon-gamma and interleukin-10-positive cells in Porphyromonas gingivalis–specific T-cell lines established from P. gingivalis–positive subjects

T-cell cytokine profiles in ten adult periodontitis and seven age-matched healthy or gingivitis subjects were determined. Porphyromonas gingivalis-specific T-cell lines were established from the peripheral blood of these individuals all of whom had past or present evidence of P. gingivalis infection. FACS analysis was used to determine the percentage of CD4- and CD8-positive cells in each line staining for cytoplasmic interleukin (IL)-4, interferon-gamma and IL-10. There were no differences in the mean percentage of IL-4-, interferon-gamma- or IL-10-positive T cells between the two groups. However, the individual profiles showed that the CD4 cells in five of the seven healthy or gingivitis lines had a higher proportion of interferon-gamma-positive cells, with two lines demonstrating higher percentages of IL-10- and/or IL-4-positive CD4 cells. Five of the ten adult periodontitis lines demonstrated either equal or higher percentages of IL-4-positive and/or IL-10-positive CD4 cells. With respect to the CD8 cells, two of the seven lines established from the healthy or gingivitis subjects and six of the ten adult periodontitis lines showed profiles with a higher percentage IL-4- and/or IL-10-positive cells. When the total T-cell contribution (CD4 plus CD8) for each T-cell line was determined from the individual CD4:CD8 ratios, only one of the healthy or gingivitis lines showed a profile with a higher proportion of IL-10-positive cells, while the results for the adult periodontitis lines were the same as indicated for the CD4 cell profiles, with five lines showing a higher percentage of IL-4- and/or IL-10-positive cells. In conclusion, this study has shown that in P. gingivalis-responsive T-cell lines established from adult periodontitis and healthy or gingivitis subjects, there was a predominant trend towards a higher percentage of interferon-gamma positive cells than either IL-4- or IL-10-positive cells. However, there were variations from this trend, although whether these variations indicate true susceptibility to progressive disease has yet to be determined.     

Interferon-γ-producing cells and inducible nitric oxide synthase-producing cells in periapical granulomas

Periapical granulomas contain a large number of T lymphocytes and monocytes/macrophages and a small number of B lymphocytes and polymorphonuclear leukocytes. Sections from eight periapical granulomas were stained by a variety of immunohistochemical methods. The vascular endothelial cells stained positively for intercellular adhesion molecule-1 and vascular cell adhesion molecule-1. Helper T cells were identified by immunostaining for CD4 and stained positively for interferon-γ (IFN-γ). However, CD4-positive T cells did not stain for interleukin-4 (IL-4). Monocytes/macrophages were identified by immunostaining for CD68 and stained positively for IL-lγ or inducible nitric oxide synthase (iNOS). IL-1β could not be detected in the same samples. No cytokine expression was observed in B cells identified by immunostaining for CD20. IFN-γ-and iNOS-positive cells could not be detected in clinically healthy periodontal ligament being used as a negative control. These results suggest that the IFN-γ-producing T cells and iNOS-positive cells may modulate the progress of disease in local inflammation sites such as in periapical granulomas.     

The proportion of interleukin-4, interferon-gamma and interleukin-10-positive cells in Porphyromonas gingivalis–specific T-cell lines established from P. gingivalis–positive subjects

T-cell cytokine profiles in ten adult periodontitis and seven age-matched healthy or gingivitis subjects were determined. Porphyromonas gingivalis–specific T-cell lines were established from the peripheral blood of these individuals all of whom had past or present evidence of P. gingivalis infection. FACS analysis was used to determine the percentage of CD4- and CD8-positive cells in each line staining for cytoplasmic interleukin (IL)-4, interferon-gamma and IL-10. There were no differences in the mean percentage of IL-4-, interferon-gamma- or IL-10-positive T cells between the two groups. However, the individual profiles showed that the CD4 cells in five of the seven healthy or gingivitis lines had a higher proportion of interferon-gamma-positive cells, with two lines demonstrating higher percentages of IL-10- and/or IL-4-positive CD4 cells. Five of the ten adult periodontitis lines demonstrated either equal or higher percentages of IL-4-positive and/or IL-10-positive CD4 cells. With respect to the CD8 cells, two of the seven lines established from the healthy or gingivitis subjects and six of the ten adult periodontitis lines showed profiles with a higher percentage IL-4- and/or IL-10-positive cells. When the total T-cell contribution (CD4 plus CD8) for each T-cell line was determined from the individual CD4:CD8 ratios, only one of the healthy or gingivitis lines showed a profile with a higher proportion of IL-10-positive cells, while the results for the adult periodontitis lines were the same as indicated for the CD4 cell profiles, with five lines showing a higher percentage of IL-4- and/or IL-10-positive cells. In conclusion, this study has shown that in P. gingivalis–responsive T-cell lines established from adult periodontitis and healthy or gingivitis subjects, there was a predominant trend towards a higher percentage of interferon-gamma positive cells than either IL-4- or IL-10-positive cells. However, there were variations from this trend, although whether these variations indicate true susceptibility to progressive disease has yet to be determined.