Roman Perez-Fernandez

1,25‐Dihydroxyvitamin D3 administration to 6‐hydroxydopamine‐lesioned rats increases glial cell line‐derived neurotrophic factor and partially restores tyrosine hydroxylase expression in substantia nigra and striatum

It has previously been demonstrated that 1,25‐dihydroxyvitamin D3 [1,25(OH)2D3] administration, whether in cell cultures or in vivo to rats, increases glial cell line‐derived neurotrophic factor (GDNF) expression levels, suggesting that this hormone may have beneficial effects in neurodegenerative disorders. This study was carried out to explore the effects of 1,25(OH)2D3 administration in a 6‐OHDA‐lesioned rat model of Parkinsons disease on GDNF and tyrosine hydroxylase (TH) expression in substantia nigra (SN) and striatum. Two groups of animals received 1,25(OH)2D3 intraperitoneally, the first group 7 days before the unilateral injection of 6‐OHDA into the medial forebrain bundle (MFB) and the second group 21 days (days 21–28) after the unilateral injection of 6‐OHDA. Animals of both groups were sacrificed on day 28. In addition, two other groups received a unilateral injection of either saline or 6‐OHDA into the MFB. Rats were killed, and the SN and striatum were then removed for GDNF and TH determination. Striatal GDNF protein expression was increased on the ipsilateral with respect to the contralateral side after 6‐OHDA injection alone as well as in 1,25(OH)2D3‐treated rats before or after 6‐OHDA administration. As expected, 6‐OHDA injection induced an ipsilateral decrease in TH‐immunopositive neuronal cell bodies and axonal terminals in the SN and striatum. However, treatment with 1,25(OH)2D3 before and after 6‐OHDA injection partially restored TH expression in SN. These data suggest that 1,25(OH)2D3 may help to prevent dopaminergic neuron damage.

Vitamin D receptor gene expression in human pituitary gland

1,25-dihydroxyvitamin D3 has important physiological effects such as calcium transport and cell growth and differentiation. These biological effects are mediated by their binding to specific intracellular receptor termed vitamin D receptor (VDR). VDR mRNA expression has been demonstrated in several tissues, but to date, there is no information about its presence in the human pituitary gland. In this report, we demonstrate VDR mRNA expression using the reverse transcription-polymerase chain reaction (RT-PCR), as well as the cellular expression of VDR by immunohistochemistry, both in the human pituitary gland. These results suggest the possibility that, like in the rat pituitary, VDR may regulate the human pituitary gene expression and hormone secretion.

1,25-Dihydroxyvitamin D3 increases striatal GDNF mRNA and protein expression in adult rats

Glial cell line-derived neurotrophic factor (GDNF) has been postulated as a possible candidate for therapeutic treatment in Parkinsons disease (PD). Recent in vitro data suggest that 1,25-dihydroxyvitamin D3 [1,25(OH)(2)D(3)] treatment may enhance GDNF mRNA expression. In the present study, using semiquantitative RT-PCR and Western blot, we have shown that 1,25(OH)(2)D(3) administration intraperitoneally, significantly increases GDNF mRNA and protein levels in the striatum of adult rats.

Association between vitamin D receptor Fokl polymorphism and serum parathyroid hormone level in patients with chronic renal failure

We investigated the relationship between vitamin D receptor (VDR) start codon polymorphism and serum levels of PTH, calcidiol, and calcium in 64 Spanish patients with chronic renal failure (CRF). An exon 2 fragment of the VDR gene was amplified by PCR, and cleaved with the restriction enzyme Fokl. The alleles were identified according to the digestion pattern obtained as F (absence of restriction site) and f (presence of restriction site). Genotype frequencies in the patient population were 54.7% FF, 28.1 % Ff and 17.2% ff, vs 46.7% FF, 43.3% Ff and 10% ff in a healthy control population. The difference between the two populations was statistically significant (p<0.01). Within the patient population, mean serum PTH level in the FF group was significantly higher (159.77±25.69 pg/ml) than in both the Ff and ff groups (106.67±19.07 and 77.55±15.85 pg/ml, respectively; p<0.05). However there were no significant differences in serum levels of calcidiol or calcium among genotypes. These results suggest that Fokl polymorphisms of the VDR gene may determine parathyroid response in CRF patients.

Deregulation of the Pit-1 transcription factor in human breast cancer cells promotes tumor growth and metastasis

The Pit-1 transcription factor (also know as POU1F1) plays a critical role in cell differentiation during organogenesis of the anterior pituitary in mammals and is a transcriptional activator for pituitary gene transcription. Increased expression of Pit-1 has been reported in human tumorigenic breast cells. Here, we found that Pit-1 overexpression or knockdown in human breast cancer cell lines induced profound phenotypic changes in the expression of proteins involved in cell proliferation, apoptosis, and invasion. Some of these protumorigenic effects of Pit-1 were mediated by upregulation of Snai1, an inductor of the epithelial-mesenchymal transition. In immunodeficient mice, Pit-1 overexpression induced tumoral growth and promoted metastasis in lung. In patients with invasive ductal carcinoma of the breast and node-positive tumor, high expression of Pit-1 was significantly correlated with Snai1 positivity. Notably, in these patients elevated expression of Pit-1 was significantly and independently associated with the occurrence of distant metastasis. These findings suggest that Pit-1 could help to make a more accurate prognosis in patients with node-positive breast cancer and may represent a new therapeutic target.

Vitamin D receptor ontogenesis in rat liver

Abstract Vitamin D through its receptor (VDR) plays a major role in bone mineral metabolism. However, VDR is also present in a variety of cell lines as well as in numerous tissues, suggesting other functions of the hormone beyond bone metabolism and mineral homeostasis. At the liver level, it has been shown that vitamin D induces numerous changes (i.e. enzyme activity level, stimulation of some metabolic pathways and stimulation of the normal liver recovery after partial hepatectomy). However, some works did not find VDR in the liver, and also used liver tissue as a negative control of VDR gene expression. In this paper, we examined fetal, neonatal and adult rat tissues for the presence of VDR using a sensitive RT-PCR technique and immunohistochemistry. We found VDR mRNA and VDR protein in rat liver at all different periods of rat life. Thus, we suggest that some of the actions of vitamin D on liver could be mediated at the genomic level through the VDR, and that the use of this tissue as a negative control of VDR gene expression is clearly inappropriate.

Expression and prognostic significance of metalloproteases and their inhibitors in luminal A and basal-like phenotypes of breast carcinoma

To analyze the expression and prognostic value of matrix metalloproteases and their tissue inhibitors in luminal A and basal-like breast carcinomas, an immunohistochemical study was performed on cancer specimens from 93 randomly selected patients with invasive primary ductal tumors of the breast (46 with and 47 without distant metastasis) and with luminal A (n = 48) (ER+, HER2-) or basal-like (HER2-, ER-, PgR-) (n = 45) lesions. Luminal B cases were too few to analyze. Specimens were also studied using tissue microarrays and specific antibodies against matrix metalloproteases 1, 2, 7, 9, 11, 13, and 14 and tissue inhibitors 1, 2, and 3. There were no significant differences in matrix metalloprotease or tissue inhibitor expression in the 2 phenotypes of tumors. In basal-like carcinomas, high scores for matrix metalloproteases 9 and 11 were significantly associated with a high distant metastasis rate. Likewise, data showed associations between matrix metalloprotease/tissue inhibitor expression by either stromal fibroblasts or mononuclear inflammatory cells and distant relapse-free survival in both tumor phenotypes. In addition, in infiltrating luminal A and basal-like tumors, we identified a prometastatic phenotype of mononuclear inflammatory cells, showing a high matrix metalloprotease/tissue inhibitor molecular profile. Expression of matrix metalloproteases and tissue inhibitors is related to the characteristics of breast tumor cells. As prognostic factors in breast carcinomas of both luminal A and basal-like phenotypes, our results point to the importance of the expression of matrix metalloproteases and tissue inhibitors by the stromal cells.

Mesenchymal Stem Cell Secretome: Toward Cell-Free Therapeutic Strategies in Regenerative Medicine

Earlier research primarily attributed the effects of mesenchymal stem cell (MSC) therapies to their capacity for local engrafting and differentiating into multiple tissue types. However, recent studies have revealed that implanted cells do not survive for long, and that the benefits of MSC therapy could be due to the vast array of bioactive factors they produce, which play an important role in the regulation of key biologic processes. Secretome derivatives, such as conditioned media or exosomes, may present considerable advantages over cells for manufacturing, storage, handling, product shelf life and their potential as a ready-to-go biologic product. Nevertheless, regulatory requirements for manufacturing and quality control will be necessary to establish the safety and efficacy profile of these products. Among MSCs, human uterine cervical stem cells (hUCESCs) may be a good candidate for obtaining secretome-derived products. hUCESCs are obtained by Pap cervical smear, which is a less invasive and painful method than those used for obtaining other MSCs (for example, from bone marrow or adipose tissue). Moreover, due to easy isolation and a high proliferative rate, it is possible to obtain large amounts of hUCESCs or secretome-derived products for research and clinical use.

Prevalence of metabolic syndrome in Galicia (NW Spain) on four alternative definitions and association with insulin resistance.

The present study evaluated the prevalence of the metabolic syndrome (MS) in a representative sample (no.=2860) of adults from the Spanish region of Galicia using the definitions of a) the World Health Organization; b) the Third Report of the National Cholesterol Education Program Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults; c) the European Group for Study of Insulin Resistance; and d) the International Diabetes Federation. In addition, we assess concordance among the different definitions, and the relationships of MS with insulin resistance (IR) as assessed by the homeostatic model assessment (HOMA-IR) index. Our results indicate a high prevalence of MS under all 4 definitions. MS prevalence was higher in men than women on all 4 definitions, and increased significantly with body mass index and age. IR was high among subjects with MS, and the HOMA index was a good discriminator of MS and non-MS on all 4 definitions, suggesting that HOMA index may be a useful predictive tool in clinical practice.

The Pit-1/Pou1f1 transcription factor regulates and correlates with prolactin expression in human breast cell lines and tumors

The transcription factor Pit-1/Pou1f1 regulates GH and prolactin (PRL) secretion in the pituitary gland. Pit-1 expression and GH regulation by Pit-1 have also been demonstrated in mammary gland. However, no data are available on the role of Pit-1 on breast PRL. To evaluate this role, several human breast cancer cell lines were transfected with either the Pit-1 expression vector or a Pit-1 small interference RNA construct, followed by PRL mRNA and protein evaluation. In addition, transient transfection of MCF-7 cells by a reporter construct containing the proximal PRL promoter, and ChIP assays were performed. Our data indicate that Pit-1 regulates mammary PRL at transcriptional level by binding to the proximal PRL promoter. We also found that Pit-1 raises cyclin D1 expression before increasing PRL levels, suggesting a PRL-independent effect of Pit-1 on cell proliferation. By using immunohistochemistry, we found a significant correlation between Pit-1 and PRL expression in 94 human breast invasive ductal carcinomas. Considering the possible role of PRL in breast cancer disorders, the function of Pit-1 in breast should be the focus of further research.