Gestational diabetes mellitus upregulates vitamin D receptor in extravillous trophoblasts and fetoplacental endothelial cells.

OBJECTIVE: Gestational diabetes mellitus (GDM) is often accompanied by low maternal vitamin D, that is, calcitriol (1,25[OH]2 vitamin D3), levels. Here, we tested the hypothesis that the placental vitamin D receptor (VDR) is regulated by calcitriol and altered in GDM with distinct changes in different placental cell types. Specifically, we aimed to localize VDR in human term placentas from normal and GDM pregnancies, to quantify its cellular expression and to study in vitro its regulation by its physiological agonist calcitriol. STUDY DESIGN: Placental tissue slides of 80 patients (40 with GDM/40 controls) were double stained for VDR and human leukocyte antigen G to identify extravillous trophoblasts (EVTs). Staining intensity was semiquantified. Quantitative real time-polymerase chain reaction and Western blotting measured VDR messenger RNA (mRNA) and protein in decidual tissue. The trophoblast cell line BeWo was used to study in vitro VDR regulation by calcitriol (0.01, 0.1, and 1 nmol/mL). RESULTS: Vitamin D receptor protein and mRNA levels are upregulated (P < .05) in EVT (1.8-fold) as well as in placental endothelium (5.8-fold) of patients with GDM. Expression of VDR is regulated by calcitriol in a bimodal manner: high doses (0.1 and 1 nmol/mL) caused downregulation, whereas the low dose (0.01 nmol/mL) resulted in VDR upregulation. CONCLUSION: Vitamin D receptor is upregulated in EVT and endothelium of GDM placentas. This could be due to low maternal vitamin D levels in patients with GDM because in vitro low calcitriol doses upregulate VDR in trophoblast cells.

Prophylactic mastectomy with immediate reconstruction combined with simultaneous laparoscopic salpingo-oophorectomy via a transmammary route: a novel surgical approach to female BRCA-mutation carriers.

INTRODUCTION: Breast reconstruction with salpingo-oophorectomy can easily be performed in patients with genetic mutations increasing the risk for mammary and ovarian carcinoma. However, many patients are skeptical about having several surgeries, as they may result in additional anesthesiological risks as well as multiple visible scars. Therefore, the purpose of this study was to evaluate the feasibility of prophylactic mastectomy and breast reconstruction combined with simultaneous transmammary salpingo-oophorectomy for BRCA carriers. MATERIALS AND METHODS: Of the six patients (1 %) who chose prophylactic mastectomy with salpingo-oophorectomy at our hospital four patients had BRCA-1 mutations, one patient had a BRCA-2 mutation and one patient had a family inheritance pattern with no mutations. All patients chose to reduce their risk for mammary and ovarian cancer by undergoing bilateral mastectomy and bilateral salpingo-oophorectomy. Prophylactic mastectomy with immediate reconstruction was performed, followed by bilateral salpingo-oophorectomy with a procedure that relies on transmammary access and reduces the number of necessary surgeries without compromising cosmetic results, surgical risks and operating time. RESULT: The mean age of the patients was 46.7 ± 1.8 years (SD). The mean operative time was 190.2 ± 13.7 min. No complications were observed during the operations. The mean intra-operative loss of blood was 363.3 ± 77.9 ml. The operative method was successful in all six cases and was performed with no complications. All of the patients were satisfied with the cosmetic results. CONCLUSION: In conclusion, prophylactic mastectomy and breast reconstruction combined with simultaneous laparoscopic salpingo-oophorectomy via transmammary access is feasible, easy to perform and provides an intriguing and novel approach to female BRCA carriers who desire operative prophylactic measures in one surgical session with no visible abdominal scars and no additional risks and complications.

Peroxisome proliferator-activated receptor-gamma (PPARγ) is down regulated in trophoblast cells of gestational diabetes mellitus (GDM) and in trophoblast tumour cells BeWo in vitro after stimulation with PPARγ agonists.

AIMS: Peroxisome proliferator-activated receptor-gamma (PPARγ) plays an important role in insulin metabolism, trophoblast differentiation and anti-inflammatory circuits. The aim of this study was to investigate the expression of PPARγ in the placenta of patients with gestational diabetes mellitus (GDM) and the regulation of PPARγ by its agonists in trophoblast tumour cells BeWo in vitro. METHODS: PPARγ expression in a total of 80 placentas (40 GDM/40 controls) was analysed by immunohistochemistry using the semi-quantitative immunoreactive score. Furthermore, a quantitative reverse transcription-polymerase chain reaction (PCR) was performed to determine the PPARγ mRNA-expression in both groups. We used a fused and a non-fused BeWo cell culture model for the stimulation with arachidonic acid and 15-Deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2). Afterwards PPARγ mRNA-expression was analysed by quantitative real-time PCR (RT-PCR) (TaqMan). RESULTS: Using immunohistochemistry we identified a decreased expression of PPARγ in the syncytiotrophoblast and the extravillous trophoblast of GDM placentas compared to normal controls. Furthermore, PPARγ mRNA-expression was reduced in GDM placentas. Stimulation of BeWo cells with arachidonic acid and 15d-PGJ2 caused a downregulation of PPARγ expression. CONCLUSION: As PPARγ is down regulated by arachidonic acid and 15d-PGJ2, the reduced PPARγ expression in GDM placentas may be due to an altered concentration of fatty acid derivates.

The liver x receptor in correlation with other nuclear receptors in spontaneous and recurrent abortions.

The liver X receptors (LXRs) have been shown to be crucially involved in maternal-fetal cholesterol transport and placentation. The aim of this study was to investigate the expression pattern and frequency of LXR under normal physiological circumstances and in spontaneous abortion and/or recurrent miscarriage. A total of 29 (12 physiologic pregnancies/10 spontaneous abortions/7 recurrent miscarriages) human pregnancies in first trimester were analysed for LXR expression. Expression changes were evaluated by immunohistochemistry for receptor and quantitative RT-PCR (TaqMan) was performed to determine the level of LXR mRNA expression. We also stained for RXR α and PPAR γ as possible heterodimers of LXR. LXR expression was downregulated in the syncytiotrophoblast of spontaneous abortion placentas compared to normal pregnancy. In recurrent miscarriage there was a trend for a downregulation. Decidua showed an even stronger downregulation in both groups. In the syncytiotrophoblast we found a positive correlation for the combination of LXR/PPAR γ in abortions and a negative correlation for LXR/RXR α . In addition, double-immunofluorescence staining showed that LXR as well as RXR α and PPAR γ are expressed by the extravillous trophoblast. Finally, RXR α and LXR showed coexpression in the same extravillous trophoblast cells. To conclude, our data show that LXR expression is decreased in miscarriage.

Bilateral prophylactic skin-reducing nipple-sparing mastectomy with immediate breast reconstruction using only a vascularized dermal-subcutaneous pedicle: technique and possible advantages.

PURPOSE: To present a new surgical technique regarding breast reconstruction after skin-reducing nipple-sparing mastectomy. METHOD: The current trend for immediate breast reconstruction after skin-reducing mastectomy mainly supports the insertion of subpectoral implants or the use of autologous breast reconstruction techniques. Herein for the first time, we present a case of bilateral prophylactic skin-reducing nipple-sparing mastectomy with immediate breast reconstruction, using only a dermal-cutaneous pedicle. RESULTS: The postoperative course was uneventful. Forty days postoperatively the aesthetic result was excellent. CONCLUSIONS: We believe that such technique in selected cases can present several advantages as low cost, reduced possibilities for complications associated to implant insertion or autologous reconstruction techniques and an easier mammography follow-up.

Detection of circulating tumour cells on mRNA levels with established breast cancer cell lines.

Circulating tumour cells were detected and quantified by real-time polymerase chain reaction (PCR) in peripheral blood, based on the fact that the expression of certain genes is upregulated in tumour tissues in comparison to surrounding blood cells. Calibration curves showing gene expression as functions of the number of tumour cells within a blood sample were prepared. Blood samples were therefore spiked with cells of breast cancer cell lines, RNA was extracted, transcribed to complementary DNA (cDNA) and used in real-time PCR reaction on the Cytokeratins (CK) 8, 18 and 19. Calibration curves were generated by Microsoft™ Excel®. Relative quantification curves of gene expression in different breast cancer cell lines showed no unitary tendencies. The oscillations in the relative quantification curves of gene expression suggested an occurrence of immunological effects, leading to an apparent agglutination of added tumour cells together with the blood cells of the sample. Thus, strategies to obtain evaluable results should be considered.

Trastuzumab clears HER2/neu-positive isolated tumor cells from bone marrow in primary breast cancer patients.

PURPOSE: Isolated tumor cells (ITC) in the bone marrow of breast cancer patients increase the risk of recurrence and decrease survival, both at primary diagnosis and during follow-up. We tested the efficacy of trastuzumab in clearing HER2/neu-positive ITC from the marrow of patients completing primary treatment. METHODS: Ten recurrence-free patients with persistent HER2/neu-positive ITC after routine adjuvant treatment received trastuzumab 6 mg/kg q3w for 12 months in a non-randomized pilot phase II interventional study. Bone marrow ITC HER2/neu status was evaluated at baseline, after treatment for 3, 6 and 12 months, and yearly thereafter, in combination with clinical follow-up. Median follow-up was 23 (15-64) months after baseline bone marrow aspiration. RESULTS: Trastuzumab for 12 months eradicated HER2/neu-positive ITC from bone marrow in all patients (P = 0.002) and significantly reduced the number of ITC-positive patients (P = 0.031). However, HER2/neu-negative ITC persisted in three patients immediately after treatment and were detected at yearly bone marrow aspiration in five patients. Two patients with ITC counts ≥5 at yearly follow-up developed metastases and one died. CONCLUSION: This is the first evidence that trastuzumab is effective in clearing HER2/neu-positive cells from bone marrow during recurrence-free follow-up in breast cancer patients. It also suggests, thanks to the antigen shift phenomenon, an important prognostic role for HER2/neu expression on marrow ITC as a real-time biopsy. However, treatment was mainly effective in patients with HER2/neu-positive ITC. Given the heterogeneity of minimal residual disease, these patients might benefit from a combination of targeted treatment approaches.