Cytoplasmic phospholipase A2 alpha overexpression in stromal cells is correlated with angiogenesis in human colorectal cancer.

In colorectal cancer, cyclooxygenase-2 (COX-2) overexpression in stromal cells induces angiogenesis through EP2 prostaglandin E2 receptor signaling. Cytoplasmic phospholipase A2 (PLA2) alpha preferentially hydrolyses arachidonic acid, which is the limiting substrate for prostaglandin production, from membrane phospholipids. We therefore investigated a possible relationship between cytoplasmic PLA2 and COX-2 overexpression in stromal cells, angiogenesis and microsatellite instability in 48 human colorectal adenocarcinomas. Cytoplasmic PLA2 and COX-2 expression in stromal cells and vascular endothelial growth factor (VEGF) expression in tumor cells were evaluated by immunohistochemistry. Microvessel density was assessed in 10 x 400 fields after CD31 staining. Microsatellite instability was evaluated by PCR and immunohistochemistry. A total of 16 tumors had microsatellite instability. We found an overexpression of cytoplasmic PLA2 in superficial stromal cells. These cells corresponded to fibroblasts and myofibroblasts. There was an association between the number of cytoplasmic PLA2 and COX-2-expressing cells (P=0.006). Cytoplasmic PLA2-positive stromal cells usually also expressed COX-2. A high number of cytoplasmic PLA2-positive stromal cells was correlated with a high microvessel density (P=0.002), a strong VEGF (P=0.01) and the absence of microsatellite instability (P=0.001). The coordinate overexpression of cytoplasmic PLA2 and COX-2 in stromal cells could lead to an important prostaglandin production. These results suggest that cytoplasmic PLA2 overexpression in these cells regulates COX-induced angiogenesis probably by providing arachidonic acid, which is the limiting factor for prostaglandin production. The lower number of cytoplasmic PLA2-positive stromal cells in carcinomas with microsatellite instability could be related to their lower microvessel density and VEGF expression.     

骨髓基质细胞促进人胚神经干细胞向神经元的分化

目的:探讨骨髓基质细胞(BMSCs)对人胚神经干细胞(NSCs)分化的影响。方法:采用机械法分离人胚NSCs,成球法进行传代培养,采用免疫荧光染色检测神经上皮干细胞蛋白(Nestin)的表达鉴定NSCs。按培养方式不同,分为NSCs自然分化组、BMSCs和NSCs直接接触共培养组及Transwell共培养组,采用免疫细胞荧光法及免疫印迹法检测各组神经元和星形胶质细胞标志物的表达。结果:在直接接触共培养组和transwell共培养组中,免疫荧光染色显示神经元标志物NSE阳性细胞率明显高于自然分化组,而星形胶质细胞标志物GFAP阳性细胞率低于自然分化组。免疫印迹检测显示Transwell共培养组中NSE表达量显著高于自然分化组,而GFAP表达量低于自然分化组。结论:BMSCs具有促进NSCs向神经元分化的作用。     

宫颈癌细胞通过分泌TSLP促进自身细胞活力

目的:探讨胸腺基质淋巴细胞生成素(TSLP)对宫颈癌细胞活力的自分泌调节作用。方法:体外培养宫颈癌细胞株HeLa和CasKi,分别收集培养24、48和72小时的培养上清,ELISA法检测培养上清中TSLP的分泌水平;流式细胞术分析HeLa和CasKi细胞表面TSLP受体(TSLPR)的表达水平;再用MTT法分别检测HeLa和CasKi细胞经人重组细胞因子TSLP(1、10和100 ng/ml)处理24、48和72小时后细胞活力的水平变化。结果:HeLa和CasKi细胞均呈时间依赖性分泌TSLP(P<0.01),且HeLa细胞在24和48小时分泌TSLP的水平高于CasKi细胞(P<0.01或P<0.05);HeLa和CasKi细胞TSLPR阳性表达率分别为23.61±1.30和27.07±2.13,前者低于后者(P<0.05);此外,10或100 ng/ml TSLP作用HeLa和CasKi细胞48小时或72小时均能上调其细胞活力(P<0.05)。结论:宫颈癌细胞可能通过TSLP自分泌作用促进自身的活力,进而利于宫颈癌细胞的生长,因而参与宫颈癌的发病和进展。     

Cytoplasmic suppression of tumor progression in reconstituted cells

This report details studies of whether mouse NIH/3T3 TG^r karyoplasts that are exposed to benzo[a]pyrene epoxide(trans) (BPDE) can progress to tumorigenicity when they are rescued with either mouse B10mtJ CAP^r tumorigenic (experiment 1) or nontumorigenic (experiment 2) cytoplasts. The mitochondrial DNA of the B10mtJ cells has restriction fragment length differences that allow distinction from the mitochondrial DNA of the NIH/3T3 cells. The reconstructed clones in experiment 1 were all tumorigenic, while those from experiment 2 were all nontumorigenic. The clones in both experiments were passaged for an equivalent time. These findings reflect the presence of factors in mouse cytoplasm capable of suppressing the tumor phenotype of NIH/3T3-BPDE treated karyoplasts when rescued at an early stage of progression.     

细胞因子诱导杀伤细胞分泌因子影响人肝癌干细胞的凋亡

BACKGROUND: Immunotherapy with autologous immune cells has been developed as a major adjuvant therapy for malignant tumors, but its mechanism of action has not been elucidated. OBJECTIVE: To investigate the relationship between cytokine-induced killer cell secretion and apoptosis in human liver cancer stem cells. METHODS: Human liver cancer stem cells, HepG2 cells, were isolated and enriched using serum-free suspension method. The peripheral blood mononuclear cells from patients with liver cancer were induced by γ-interferon, CD3 monoclonal antibody and recombinant human interleukin-2 to form killer cells. Passage 1 liver cancer stem cells were divided into control group (culture alone) and experimental group (co-culture of cytokines-induced killer cells and human liver cancer stem cells). At 48 hours after culture, apoptosis in human liver cancer stem cells was detected using flow cytometry, and expression of caspase-3 mRNA and protein was detected using RT-PCR and western blot, respectively. RESULTS AND CONCLUSION: The apoptotic rate in the control group was significantly lower than that in the experimental group (P < 0.05). The expressions of caspase-3 at mRNA and protein levels were both higher in the experimental group than the control group (P < 0.05). Experimental findings show that cytokines-induced killer cells can significantly promote apoptosis in human liver cancer stem cells, and up-regulate the caspase-3 mRNA and protein expressions dramatically.  中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程      

Swelling-activated taurine and K+ transport in human cervical cancer cells: association with cell cycle progression

The aim of this study was to investigate swelling-activated taurine and K+ transport in human cervical cancer cells under various culture conditions, testing the hypothesis that the progression of cell cycle was accompanied by differential activities of swelling-activated transport pathways. Aphidicolin, an inhibitor of deoxyribonucleic acid (DNA) synthesis, was used to synchronize the cell cycle. The distribution of cell cycle stage was determined by fluorescence-activated cell sorting (FACS). Hypotonicity activated taurine efflux, which was sensitive to tamoxifen and 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB). Cell swelling also induced both Cl- -dependent and -independent K+ (86Rb+) efflux, presumably mediated by KCl cotransport (KCC) and Ca2+ -activated K+ channels, respectively. Cell cycle arrest in G0/G1 was accompanied by a remarkable decrease in the rate constant for swelling-activated taurine efflux, from 0.20+/-0.007 to 0.026+/-0.002 min(-1) (n=6). The activity of swelling-activated taurine efflux recovered progressively on re-entry into the cell cycle. After removal of aphidicolin and culture with 10% fetal calf serum for 10 h, the rate constant increased significantly from 0.026+/-0.002 to 0.093+/-0.002 min(-1) (n=6). After 24 h release from aphidicolin, the efflux rate constant had increased further to 0.195+/-0.006 min(-1) (n=6), a value not significantly different from that in normally proliferating cells. The differential activities of swelling-activated taurine transport matched well with our previous study showing a volume-sensitive anion channel associated with cell cycle progression. In contrast to the differential activities of swelling-activated taurine transport, swelling-activated K+ (86Rb+) transport was independent of the progression of cell cycle. Most importantly, pharmacological blockade of swelling-activated taurine efflux by tamoxifen or NPPB caused proliferating cervical cancer cells to arrest in G0/G1, suggesting that the activity of this efflux was associated with G1/S checkpoint progression. This study provides new and important information on the functional significance of swelling-activated transport system in the regulation of cell cycle clock of human cervical cancer cells.     

Early stromal invasion in cervical cancer: immunocytochemical changes occurring in infiltrating neoplastic cells

Early Stromal Invasion (ESI) in cervical cancer progression should be considered as a separate histological diagnostic category for its morphological characters very different from those of both carcinoma in situ (CIS) and microcarcinoma (MIC). To have some more microscopical details on these differences we performed immunocytochemical investigation addressed to evaluate, in cervical cancer malignancy progression, the evolutionary changes in the expression of some proteins involved in cell differentiation and cell cycle regulation. The results provide data improving the knowledge about ESI and supporting, with objective proofs, the nosological autonomy of ESI, with respect to CIS and MIC.     

Cytoplasmic overexpression of ALCAM is prognostic of disease progression in breast cancer

BACKGROUND: Activated leucocyte cell adhesion molecule (ALCAM, CD166) is a cell surface member of the immunoglobulin superfamily. ALCAM expression has prognostic relevance in prostate and colon cancer. OBJECTIVE: To evaluate ALCAM protein expression in breast cancer by immunohistochemistry and to correlate expression levels with clinicopathological data. METHODS: 162 primary breast carcinomas with a mean clinical follow up time of 53 months were immunostained using a monoclonal ALCAM antibody. The staining was evaluated as an immunoreactive score (IRS) and grouped into low v high for both membranous and cytoplasmic staining. RESULTS: Intraductal and invasive carcinomas showed a higher ALCAM expression (median IRS 4 and 6 respectively) than normal breast tissue (IRS 2). In univariate survival analyses a significant association of high cytoplasmic ALCAM expression with shortened patient disease-free survival (mean (SD) five year non-progression rate, 69.4 (4.6)% v 49.4 (11.1)%, p = 0.0142) was found. In multivariate analyses of disease-free survival times, high cytoplasmic ALCAM expression (relative risk (RR) = 2.086, p = 0.026) and nodal status (RR = 2.246, p = 0.035) were significantly associated with earlier disease progression, whereas tumour grading (RR = 1.6, p = 0.052) was of borderline significance. CONCLUSIONS: The data suggest that strong cytoplasmic ALCAM expression in primary breast cancer, as detected by immunohistochemistry, might be a new marker for a more aggressive breast cancer biology.     

Xenogeneic chondrocytes promote stable subcutaneous chondrogenesis of bone marrow-derived stromal cells

The local microenvironment may change the ultimate fate of engineered cartilage differentiated from bone marrow stromal cells (BMSCs) after subcutaneous implantation. Chondrogenically differentiated BMSCs directed by growth factors or low-intensity ultrasound are apt to fibrose or vascularize in the subcutaneous environment, while BMSCs implanted in articular cartilage defects can form stable cartilage. We hypothesized that chondrocytes would provide an ideal chondrogenic environment, and thus promote the maintenance of the chondrocytic phenotype in ectopia. To test this hypothesis, we developed a new method to promote chondrocyte development from BMSCs in a chondrogenic environment produced by xenogeneic chondrocytes and compared the subcutaneous chondrogenesis of BMSCs mediated by xenogeneic chondrocytes with that produced by growth factors. These results indicate that subcutaneous chondrogenesis of BMSCs directed by xenogeneic chondrocytes is more effective than that induced by growth factors. BMSCs induced by xenogeneic chondrocytes formed relatively mature cartilage before or after implantation, following 4 weeks of culture, which reduced the induction time in?vitro and led to maintenance of a stable cartilage phenotype after subcutaneous implantation.     

Decidual stromal cells recruit Th 17 cells into decidua to promote proliferation and invasion of human trophoblast cells by secreting IL-17

T helper 17 （Th17） cells have both regulatory and protective roles in physiological conditions. The Th17 subset and the cytokine interleukin-17A （IL-17A） have been implicated in the pathogenesis of certain autoimmune diseases, several types of cancer and allograft rejection. However, the role of Th17 cells at the maternal/fetal interface remains unknown. Here, we demonstrate that Th17 cells are present in decidua and are increased in the peripheral blood of 10 clinically normal pregnancies based on intracellular cytokine analysis. Our results suggest a potential role of Th17 cells in sustaining pregnancy in humans. Furthermore, we demonstrate that decidual stromal cells （DSCs） but not trophoblast cells recruit peripheral Th17 cells into the decidua by secreting CCL2. The recruited Th17 cells promote proliferation and invasion and inhibit the apoptosis of human trophoblast cells by secreting IL-17 during the first trimester of pregnancy. These findings indicate a novel role for Th17 cells in controlling the maternal-fetal relationship and placenta development.     

Interleukin‐17 and interleukin‐22 promote tumor progression in human nonmelanoma skin cancer

Interleukin‐17 (IL‐17) and IL‐22 have been reported to play critical roles in autoimmunity and inflammation but information about their role in cancer is limited. In this study, we investigated the role of IL‐17 and IL‐22 in the progression of human skin basal‐cell carcinoma (BCC) and squamous‐cell carcinoma (SCC). We found that both tumor types are infiltrated with an high number of IL‐17⁺ and IL‐22⁺ T lymphocytes, as demonstrated by immunohistochemistry and by FACS analysis performed on peritumoral T‐cell lines isolated from skin biopsies. In vitro studies demonstrated that proliferation and migration of the BCC‐ and SCC‐cell lines M77015 and CAL27 were increased by IL‐17 and IL‐22. Moreover, IL‐17, alone or in combination with TNF‐α, was able to induce the production of two cytokines important for tumor progression, IL‐6 and IL‐8, in CAL27. We also showed that IL‐17 upregulated NF‐κB signaling, while IL‐22 activated the STAT3 pathway and the antiapoptotic AKT protein in M77015 and CAL27. Finally, in vivo experiments demonstrated that IL‐17 and IL‐22 enhanced tumor growth in nude mice injected with CAL27. Altogether, our findings indicate that high levels of IL‐22 and IL‐17 in the BCC and SCC microenvironment promote tumor progression.     

Changes in retinoblastoma gene expression during cervical cancer progression

The role of tumour suppressor genes in the development of human cancers has been studied extensively. In viral carcinogenesis, the inactivation of suppressor proteins such as retinoblastoma (pRb) and p53, and cellular oncogenes overexpression, such as c-myc, has been the subject of a number of investigations. In uterine-cervix carcinomas, where high-risk human papillomavirus (HPV) plays an important role, pRb and p53 are inactivated by E7 and E6 viral oncoproteins, respectively. However, little is known about the in situ expression of some of these proteins in pre-malignant and malignant cervical tissues. On the other hand, it has also been demonstrated that c-myc is involved in cervical carcinogenesis, and that pRb participates in the control of c-myc gene expression. By using immunostaining techniques, we investigated pRb immunodetection pattern in normal tissues, squamous intraepithelial lesions (SILs) and invasive carcinomas from the uterine cervix. Our data show low pRb detection in both normal cervical tissue and invasive lesions, but a higher expression in SILs. C-Myc protein was observed in most of the cellular nuclei of the invasive lesions, while in SILs was low. These findings indicate a heterogeneous pRb immunostaining during the different stages of cervical carcinogenesis, and suggest that this staining pattern could be a common feature implicated in the pathogenesis of uterine-cervix carcinoma.     

力达霉素诱导人宫颈癌Caski 细胞发生免疫原性细胞凋亡

目的:探讨抗肿瘤药物力达霉素能否抑制人宫颈癌Caski 细胞增殖并且诱导Caski 细胞发生免疫原性细胞凋亡。方法:MTT 比色法检测Caski 细胞增殖;Annexin V-FITC/ PI 双染流式细胞术检测Caski 细胞凋亡。Western blot 分析细胞中Bax 和Bcl-2 的表达;免疫荧光流式细胞术检测细胞凋亡后细胞膜上钙网蛋白的表达。结果:力达霉素抑制Caski 细胞的增殖,具有时间和剂量依赖性;5 g/ L 的力达霉素作用Caski 细胞48 h 后,肿瘤细胞凋亡率为11.5%;Bax 表达量显著增加,而Bcl-2 含量明显减少(P<0.05);细胞膜表面钙网蛋白表达高达67.2%,显著高于对照组的2.31%。结论:力达霉素能够有效抑制人宫颈癌Caski 细胞增殖,其作用机制与线粒体途径诱导细胞凋亡相关,同时促使钙网蛋白转移到Caski 细胞膜表面表达,可能具有诱导肿瘤细胞发生免疫原性细胞凋亡的能力。     

CRH通过CRHR1激活cAMP/PKA通路调节宫颈癌细胞的免疫逃逸和促进宫颈癌生长

目的 探讨促肾上腺皮质激素释放激素(corticotropin-releasing hormone,CRH)及其受体(corticotropin-releasing hormone receptor 1,CRHR1)影响宫颈癌免疫逃逸和生长的途径.方法 采用实时荧光定量聚合酶链反应(quantitative real-...     

Mesenchymal stromal cells in cancer: a review of their immunomodulatory functions and dual effects on tumor progression

Mesenchymal stem or stromal cells (MSCs) are pluripotent cells implicated in a broad range of physiological events, including organogenesis and maintenance of tissue homeostasis as well as tissue regeneration and repair. Because their current definition is somewhat loose – based primarily on their ability to differentiate into a variety of mesenchymal tissues, adhere to plastic, and express, or lack, a handful of cell surface markers – MSCs likely encompass several subpopulations, which may have diverse properties. Their diversity may explain, at least in part, the pleiotropic functions that they display in different physiological and pathological settings. In the context of tissue injury, MSCs can respectively promote and attenuate inflammation during the early and late phases of tissue repair. They may thereby act as sensors of the inflammatory response and secrete mediators that boost or temper the response as required by the stage of the reparatory and regenerative process. MSCs are also implicated in regulating tumor development, in which they are increasingly recognized to play a complex role. Thus, MSCs can both promote and constrain tumor progression by directly affecting tumor cells via secreted mediators and cell–cell interactions and by modulating the innate and adaptive immune response. This review summarizes our current understanding of MSC involvement in tumor development and highlights the mechanistic underpinnings of their implication in tumor growth and progression. © 2020 Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.     

Immunohistochemical study of human placental stromal cells

An immunohistochemical study of stromal cells in human placental villi was made using various polyclonal and monoclonal antibodies. This study demonstrated that almost all villous stromal cells expressed HLA-ABC, which is indicative of class I major histocompatibility complex, and vimentin, which is a mesenchymal marker, through the entire period of pregnancy. Some of these stromal cells were considered to be fetal macrophages having HLA-DR, which is a determinant of class II major histocompatibility complex, particularly in the second and third trimesters. Such macrophages also expressed CD-4 (Leu-3a and -3b) and 2H4, which are the cell membrane determinants of suppressor-inducer T lymphocyte; CD-2 (Leu-5b), which is a marker of pan-T cell; Leu-M3, Leu-M5 and Mac-1, which are the markers of monocyte and macrophage lineage; leukocyte common antigen, which is a marker of bone marrow derived cell; and alpha-antichymotrypsin, which is a glycoprotein associated with macrophages. Morphologically, villous macrophages consisted of heterogeneous phenotypes such as classic Hofbauer cells, fibroblast-like spindle cells with long cytoplasmic processes, and dendritic-shaped cells. These may have more complex features than previously considered and may have a greater initiating role in immunologic interactions between mother and fetus when compared with the very sparsely distributed T or B lymphocytes.