Combined therapy with disintegrin and melphalan as a new strategy in inhibition of endometrial cancer cell line (Ishikawa) growth.

Endometrial cancer is one of the most frequently diagnosed cancer in females with prevalence of 22 in 100,000 women. The etiology of the cancer remains unclear. Despite significant progress towards understanding the patho-mechanism of the disease, effective treatment is still lacking. The results of the study suggest that combined treatment of Ishikawa cells for 24 h with disintegrin and then for 24 h with melphalan severely inhibits important biological functions of the cells. We showed that such strategy have a potent cytotoxic effect. The mechanism of process undergoes probably through inhibition of integrin - dependent signaling. In this study we shown down regulation of Shc and FAK proteins in cells treated with echistatin and melphalan. It suggests that signaling pathways that involve Shc and FAK participation may represent target for antineoplastic strategy. The functional significance of the combined treatment of Ishikwa cells with echistatin and melphalan was found at the level of collagen biosynthesis. Decreased biosynthesis of collagen in extracellular matrix may suppress cell growth and induce apoptosis. The treatment with echistatin and melphalan also showed decreased expression of IGF receptor in comparison to the cells treated with both compounds separately. The data presented suggest that combined therapy with disintegrin - echistatin and alkyalting drug - mephalan may represent a new approach to more effective and safe cancer therapy

Mechanism of collagen biosynthesis up-regulation in cultured leiomyoma cells.

Uterine leiomyoma is the most common tumour in women with a reported incidence of 25-30%. The tumors are benign, composed of smooth muscle cells with variable amount of collagen - rich fibrous tissue. It is well established that accumulation of extracellular matrix in leiomyoma is key feature of tissue fibrosis. However, the pathogenesis of leiomyoma is still unclear. The aim of this study was to evaluate the metabolism of collagen in cultured leiomyoma cells and in control myometrium cells. The effect of estradiol, selective modulators of estrogen receptors (raloxifene, tamoxifen) and estrogen receptor down regulator (ICI 182.780) on collagen biosynthesis (measured by 5-[3H]-proline incorporation assay and measurement of prolidase activity) and collagen degradation (measured by metalloproteinase activity assay) was studied. It was found that collagen biosynthesis is strongly stimulated by low doses of estradiol (5 nM) in leiomyoma cells while it is not changed in control myometrium cells. An increase in estradiol concentration to 10 nM results in drastic decrease of this process both in leiomyoma as well as control cells. Although raloxifene and tamoxifen only slightly affected collagen biosynthesis in control myometrium cells, they significantly inhibited the process in leiomyoma cells. There was no coordinate correlation between collagen biosysignificantly inhibited the process in leiomyoma cells. There was no coordinate correlation between collagen biosynthesis and prolidase activity suggesting that regulation of this process may take place at transcriptional level. Both estrogen and SERMs were found to inhibit MMP-2 in leiomyoma as well as in control myometrium cells. The data suggest that stimulatory action of estrogen on collagen biosynthesis and inhibitory effect on MMP-2 activity in uterine leiomyoma may contribute to accumulation of this protein in ECM of this tissue

Applying next-generation sequencing platforms for pharmacogenomic testing in clinical practice

Pharmacogenomics (PGx) studies the use of genetic data to optimize drug therapy. Numerous clinical centers have commenced implementing pharmacogenetic tests in clinical routines. Next-generation sequencing (NGS) technologies are emerging as a more comprehensive and time- and cost-effective approach in PGx. This review presents the main considerations for applying NGS in guiding drug treatment in clinical practice. It discusses both the advantages and the challenges of implementing NGS-based tests in PGx. Moreover, the limitations of each NGS platform are revealed, and the solutions for setting up and management of these technologies in clinical practice are addressed.Personalised Therapeutic

Estrogen receptor beta participate in the regulation of metabolizm of extracellular matrix in estrogen alpha negative breast cancer.

The biology of breast cancer is closely releted to sex steroid hormones. Estrogen receptor beta is overexpressed in around 70% breast cancer cases, referrd to as "ER positive". Estrogens bind to estrogen receptor and stimulate the transcription of genes involved in control of cell proliferation. Moreover, estrogens may induce growth factors and components of extracellular matrix and interact with them in a complex manner. Extracellular matrix and integrins play an important role in cell functions and their aberrant expressions are implicated in breast cancer development, invasion and metastasis. ER beta is certainly associated with more differentiated tumors, while evidence of role of ER beta is controversial. The highly invasive breast cancer ER beta negative cell line MDA-MB 231 can be the model of exam the role of ER beta in breast cancer. The aim of this study was to examine the role of activation of ER beta on the metabolism of the extracellular matrix and the expression of beta-1 integrin in the breast cancer cell line MDA-MB 231. The cells were exposed on the estradiol, tamoxifen, raloxifen and genisteina in dose dependent concentrations. To determine the relative rate of collagen syntesis we measured the time-dependent reduction of collagen-bound radioactivity after pulse-chase labeling with [3 H] prolina by Peterkofsky methods. The expression of beta-1 integrin was determine by Western blot analysis. The activity of MMP2 and 9 were measured using gelatin zymography with an image analysis system. Our data suggest on the role of estrogen receptor beta on the metabolism of extracellular matrix in the breast cancer line MDA - MB 231. Estradiol and SERMs regulate the expression of ECM proteins: collagen, integrins and enhance activity of metaloproteinases 2 and 9

Functional Consequences of Intracellular Proline Levels Manipulation Affecting PRODH/POX-Dependent Pro-Apoptotic Pathways in a Novel in Vitro Cell Culture Model

Background/Aims: The effect of impaired intracellular proline availability for proline dehydrogenase/proline oxidase (PRODH/POX)-dependent apoptosis was studied. Methods: We generated a constitutively knocked-down PRODH/POX MCF-7 breast cancer cell line (MCF-7(shPRODH/POX)) as a model to analyze the functional consequences of impaired intracellular proline levels. We have used inhibitor of proline utilization in collagen biosynthesis, 2-metoxyestradiol (MOE), inhibitor of prolidase that generate proline, rapamycin (Rap) and glycyl-proline (GlyPro), substrate for prolidase. Collagen and DNA biosynthesis were evaluated by radiometric assays. Cell viability was determined using Nucleo-Counter NC-3000. The activity of prolidase was determined by colorimetric assay. Expression of proteins was assessed by Western blot and immunofluorescence bioimaging. Concentration of proline was analyzed by liquid chromatography with mass spectrometry. Results: PRODH/POX knockdown decreased DNA and collagen biosynthesis, whereas increased prolidase activity and intracellular proline level in MCF-7(shPRODH/POX) cells. All studied compounds decreased cell viability in MCF-7 and MCF-7(shPRODH/POX) cells. DNA biosynthesis was similarly inhibited by Rap and MOE in both cell lines, but GlyPro inhibited the process only in MCF-7(shPRODH/POX) and MOE+GlyPro only in MCF-7 cells. All the compounds inhibited collagen biosynthesis, increased prolidase activity and cytoplasmic proline level in MCF-7(shPRODH/POX) cells and contributed to the induction of pro-survival mode only in MCF-7(shPRODH/POX) cells. In contrast, all studied compounds upregulated expression of pro-apoptotic protein only in MCF-7 cells. Conclusion: PRODH/POX was confirmed as a driver of apoptosis and proved the eligibility of MCF-7(shPRODH/POX) cell line as a highly effective model to elucidate the different mechanisms underlying proline utilization or generation in PRODH/POX-dependent pro-apoptotic pathways. (C) 2017 The Author(s) Published by S. Karger AG, Base

Serum Bisphenol A Level in Boys with Cryptorchidism: A Step to Male Infertility?

Cryptorchidism is the most common congenital birth defect in boys and affects about 2–4% full-term male neonates. Its etiology is multifactorial. Purpose. To evaluate the serum bisphenol A (BPA) levels in boys with cryptorchidism and healthy boys and to assess the risk of environmental exposure to BPA using the authors’ questionnaire. The data were acquired from a study on boys with cryptorchidism (n=98) and a control group (n=57). Prior to surgery, all patients had BPA serum levels evaluated. The size, position, rigidity of the testis, and abnormality of the epididymis of the undescended testis were assessed. Parents also completed a questionnaire on the risks of exposure to BPA in everyday life. Results. The testes in both groups were similar in size. The turgor of the undescended testis in the group of boys with cryptorchidism was decreased. Free serum BPA level in cryptorchid boys and in the control group was not statistically significant (p>0.05). The conjugated serum BPA level in cryptorchid boys and in the control group was statistically significant (p≤0.05). Total serum BPA level in cryptorchid boys and in the control group was statistically significant (p<0.05). Serum total BPA level was related with a positive answer about problems with conception (p<0.02). Conclusion. Our study indicated that high serum BPA was associated with cryptorchidism

The 18 kDa Translocator Protein (Peripheral Benzodiazepine Receptor) Expression in the Bone of Normal, Osteoprotegerin or Low Calcium Diet Treated Mice

The presence of the translocator protein (TSPO), previously named as the mitochondrial or peripheral benzodiazepine receptor, in bone cells was studied in vitro and in situ using RT-qPCR, and receptor autoradiography using the selective TSPO ligand PK11195