Effect of neoadjuvant chemotherapy on stromal CD10 antigens in breast cancer - A preliminary study.

Introduction: CD10 is a zinc-dependent peptidase (metalloproteinase). Stromal CD10 expression in breast cancer correlates with poor prognosis, oestrogen receptor negativity and higher grade. CD10 may be a potential target of new cancer therapies as it is involved in cleavage of doxorubicin. Aim: To evaluate the effect of neo-adjuvant anthracycline-based chemotherapy on status of stromal CD10 antigens in breast cancer. Materials and Methods: Patients with invasive breast cancer scheduled for anthracycline-based neo-adjuvant chemotherapy were included in the study. Tumor stromal CD10 expression was estimated before and after 3 cycles of chemotherapy, and change in its status was correlated with clinical response to chemotherapy. Results: 16 out of the 29 patients had strong CD10 expression; in these 16 patients, 14 (87.5%) were hormone receptor negative, and 14 (87.5%) had HER-2/neu overexpression. Stromal CD10 expression remained same in 13 out of 29 cases (44.83%) after chemotherapy. There was a change in CD10 expression in the remaining 16 cases (55.17%); in 13 cases (44.83%) it decreased from its pre-chemotherapy status, while its expression increased in 3 cases (10.34%). In cases of complete and partial clinical response, there was no increase in CD10 expression. Where CD10 expression had increased after chemotherapy, there was either a minor response or no response to chemotherapy. In 13 cases where CD10 expression had decreased, 12 cases had a clinical response to chemotherapy. Conclusions: Strong CD10 expression correlates with hormone receptor negativity and HER-2/neu overexpression. Stromal CD10 expression in breast cancer is not static and changes with neo-adjuvant anthracycline-based chemotherapy. A stable or decrease in CD10 expression correlates with complete or partial clinical response, while an increase in CD10 expression appears to correlate with poor clinical response. A larger series is required to determine the clinical significance of these changes. As stromal CD10 expression and its change with chemotherapy may have a prognostic significance, they should be documented in breast cancer patients before and after chemotherapy.

Adiponectin effect on the viability of human endometrial stromal cells and mRNA expression of adiponectin receptors

Adiponectin is one of the most important adipokines secreted from fatty tissue that has a direct inhibitory effect on the development of cancer cells. Adiponectin plays an important role in human reproduction system and fertility of women. Adiponectin concentration decreases in women with endometriosis and endometrial cancer. The aim of the present study was to investigate the effect of adiponectin on human endometrial stromal cell [HESC] viability as well as mRNA expression of Adipo R1 and Adipo R2 receptors. In this experimental study, eight endometrial biopsies were taken and stromal cells were separated by enzymatic digestion and cell filtrations. Stromal cells of each biopsy were divided into four groups: control, 10, 100, and 200 ng/ml adiponectin concentrations. The effect of adiponectin on viability of the normal HESCs was studied by trypan blue staining and the relative expression levels of Adipo R1 and R2 were analyzed by semi-quantitative reverse transcription polymerase chain reaction [RT-PCR]. Data were analyzed by one way ANOVA and unpaired student's t test and p<0.05 was considered significant. Adiponectin decreased viability of normal human endometrial stromal cells in a dose and time dependent manner. Expression of Adipo R1 and Adipo R2 receptors did not change in the presence of adiponectin. Adiponectin can directly influence the viability of HESCs and decrease their viability, but it didn't change expression of adiponectin receptors

In vitro action of leukemia inhibitory factor (LIF) on mid-secretory stage endometrial stromal cells collected from hormone-simulated, ovariectomized monkey and maintained in three-dimensional primary culture.

Leukemia inhibitory factor (LIF) is a pleiotropic cytokine that is known to play an important role in blastocyst implantation. The putative action of LIF in the regulation of uterine function has been examined using mid-secretory stage monkey endometrial stromal cells cultured on rat-tail collagen type I and treated with recombinant human LIF (rhLIF) or immunoneutralized LIF (in LIF) under serum-free condition. Long-term ovariectomized rhesus monkeys (n=8) underwent simulation of their menstrual cycles with steroid hormones and endometrial tissue samples were collected on cycle day 18; stromal cells were isolated and grown in primary culture on three-dimensional collagen matrix. Significant decline in cellular protein synthesis (P < 0.01) and cell proliferation index (P < 0.05) was observed in cells with increasing doses (0-1000 ng/ml) of rhLIF under serum-free in vitro condition. JAK1 expression in cultured cells increased (P < 0.01) in response to rhLIF as revealed from Western blot and confocal laser scanning microscopic examination, STAT1 and STAT2 expressions were unchanged, while pSTAT3 expression increased (P < 0.01) with increased concentration of rhLIF in culture medium. Autophosphorylation of JAK1 in endometrial stromal cells showed no change with increasing concentration (0.01 to 100 ng/ml) of rhLIF in vitro, but significant (P < 0.05) increase was observed with the time of exposure to rhLIF. Immunoneutralization of LIF or no addition of rhLIF to cultured cells led to significant (P < 0.01) increase in stromal cell proliferation index and significant (P < 0.01) decrease in the level of JAK1 and its autophosphorylation as compared to cells exposed to rhLIF alone. From the present set of experiments we conclude that rhLIF affects the physiological behaviour of monkey mid secretory stage endometrial stromal cells in vitro via the JAK-STAT signaling pathway.

Increased smooth muscle actin expression from bone marrow stromal cells under retinoic acid treatment: an attempt for autologous blood vessel tissue engineering.

Vascular replacement in vital organs is sometimes necessary for human life for example because of atherosclerosis. Blood vessel tissue engineering is applied for autologous transplantations to avoid graft rejections. Stem cells are used for blood vessel tissue engineering because they are the origin of smooth muscle cells, endothelial cells and fibroblasts. This paper shows that bone marrow stromal cells (BMSCs) can be induced to differentiate into the early stage of smooth muscle cells by using 0.01 microM retinoic acid. The differentiation of BMSCs to smooth muscle cells was detected by the expression of smooth muscle alpha actin (SM alpha-actin), the earliest smooth muscle cell marker. The SM alpha-actin marker expression was demonstrated using indirect immunofluorescence technique and Western blot analysis. The induction of BMSC to form early stages of smooth muscle cells in this study is appropriate for blood vessel tissue engineering because the early stage smooth muscle cells may be stimulated to develop vascular walls with endothelial cells using a co-culture system.

Osteoblast differentiation of human bone marrow cells under continuous and discontinuous treatment with dexamethasone

A dexametasona (Dex) induz diferenciação osteoblástica em diversos modelos de cultura de células. Este estudo investigou o efeito do tratamento contínuo e descontínuo com Dex sobre a diferenciação de células de medula óssea humana (BMSC). Células da cultura primária e da primeira passagem foram cultivadas em meio de cultura com e sem Dex 10-7 M (37ºC e 5% CO2 / 95% ar atmosférico). Aos 7, 14 e 21 dias, os seguintes parâmetros foram avaliados: proliferação e viabilidade celulares, conteúdo de proteína total, atividade de fosfatase alcalina (ALP) e formação de matriz mineralizada. Os dados foram comparados por análise de variância a dois critérios. A Dex não afetou a viabilidade celular e o conteúdo de proteína total, mas reduziu o número de células. A atividade de ALP e a formação de matriz mineralizada foram aumentadas quando apenas a primeira passagem ou cultura primária e primeira passagem foram tratadas com Dex, em comparação aos grupos que não tiveram contato com Dex após a primeira passagem. Estes resultados indicam que, para BMSC humanas, a presença contínua de Dex não parece ser necessária para o desenvolvimento do fenótipo osteoblástico. Contudo, a Dex deve estar presente após a primeira passagem para permitir a diferenciação osteoblástica expressa por proliferação celular reduzida e aumento da atividade de ALP e da formação de matriz mineralizada.

Tissue alterations in the Guinea pig lateral prostate following antiandrogen flutamide therapy

The flutamide antiandrogenic effects on the Guinea pig male prostate morphology in puberal, post-puberal and adult ages were evaluated in the present study. Daily-treated group animals received flutamide subcutaneous injection at a dose of 10 mg/Kg body weight for 10 days. The control group animals received a pharmacological vehicle under the same conditions. The lateral prostate was removed, fixed and processed for light and transmission electron microscopy. The results revealed an increase of the acinus diameter in the treated puberal animals and straitness in the stromal compartment around the acini. The epithelial cells exhibited cubic phenotype. In the post-puberal and adult animals, a decrease of the acinus diameter was observed, as well as an increase of the smooth muscle layer and presence of the folds at epithelium. The ultrastructural evaluation of the secretory cells in the treated group demonstrated endomembrane enlargement, mainly in the rough endoplasmic reticulum and Golgi apparatus. In addition, a decrease of the microvilli and alterations in the distribution patterns and density of the stromal fibrillar components were observed. In conclusion, the flutamide treatment exerts tissue effects on the lateral prostate, promoting stroma/epithelium alterations.

Interaction of TGF-beta1 and rhBMP-2 on Human Bone Marrow Stromal Cells Cultured in Collagen Gel Matrix

Transforming growth factor-beta1 (TGF-beta1) and bone morphogenetic protein-2 (BMP-2) are abundant proteins in the bone matrix. However, their interaction in controlling osteoblast differentiation is not clearly understood. In this study, HBMSCs were cultured in collagen gel matrix with different condition of exogenous rhBMP-2 and TGF-beta1 in order to determine the interaction of BMP-2 and TGF-beta1 on human bone marrow stromal cells (HBMSCs) differentiation. The cultured cells were analyzed for cell proliferation, alkaline phophatase (ALP) activity and mineralization stainning with Von-Kossa. The cells treated with TGF-beta1 exhibited a higher rate of cell growth than those without. However, the cells cultured in collagen gel matrix showed a lower rate of cell growth than the cells cultured in a monolayer. To investigate the effects of both cytokines on osteoblast differentiation, the cells were treated with 0, 1, 5, 10 ng/ml of TGF-beta1 for 2 days. This was followed by culturing with 0, 1, 5, and 10 ng/ml of TGF-beta1 and 100 ng/ml of rhBMP-2 together for 3 days with the alkaline phosphatase (ALP) activity measured. The cells treated with 1 ng/ml of TGF-beta1 responded efficiently to rhBMP-2 and expressed ALP activity with a level equivalent to that exhibited by cells that were not treated with TGF-beta1. The cells treated with 5 and 10 ng/ml of TGF-beta1 showed a dramatic decrease in ALP activity. The cells treated with 10ng/ml of TGF-beta1 followed by rhBMP-2 alone exhibited an intermediate ALP activity. The cells treated with 100 ng/ml of rhBMP-2 demonstrated Von-Kossa positive solid deposits after 3 weeks, while there were few Von-Kossa positive solid deposits when the cells treated with 10 ng/ml of TGF-beta1. These results show that TGF-beta1 inhibits the effects of rhBMP-2 on the osteoblast differentiation of HBMSCs in a dose dependant manner. Furthermore, the effects of TGF-beta1 on HBMSCs are reversible. This suggest that TGF-beta1 and rhBMP-2 are coordinately controlled during the osteoblast differentiation of HMBSCs.

Tamoxifen increases the proliferation of human endometrial stromal cells in in vitro. A model for evaluation of endometrial hyperplasia.

Tamoxifen given for breast cancer therapy, has a complex and an unclear action on the endometrium. A large number of literatures has attributed the proliferous changes in the endometrium caused by tamoxifen (Tam). No report has appeared on the endometrial cellular changes induced by Tam. The present study shows a significant (P < 0.001) increase in the proliferative activity due to Tam in endometrial stromal cells over control and estradiol (E2). This in vitro model is useful for the study of the hyperplasic effect of Tam at the cellular level.