ABSTRACT
Label-free quantitative analysis of the cardiovascular structure and function in wild fish is highly important and still unsolved problem. We developed a robust algorithm for bright field microscopy data processing, which allows reliable detection and quantification of circulatory system across the whole animal. We verified this technique by in vivo experiments on broad whitefish (Coregonus nasus) embryos. Calculated morphofunctional features of heart and vessels are quite specific due to low temperature embryonic development of this fish, and correspond well to the reference data obtained by ex vivo techniques. This research highlights the feasibility of non-invasive cardiovascular imaging and measurements valuable for behavioral, environmental, toxicological, drug discovery and many other studies.
Subject(s)
Cardiovascular System , Salmonidae , Animals , Embryonic DevelopmentABSTRACT
OBJECTIVE: To identify the structural and immunohistochemical features of the placentas and the placental sites after in vitro fertilization (IVF) with a donor egg (surrogate motherhood). SUBJECT AND METHODS: Morphological and immunohistochemical studies were performed on the placental (a placental disk) and placental bed materials obtained after caesarean delivery. The investigation enrolled 26 patients whose pregnancy occurred with IVF with a donor egg according to the surrogacy (IVF-S) program. A comparison group included 13 patients whose pregnancy occurred after IVF with their own eggs. An immunohistochemical study was conducted on paraffin sections made from biopsy material; mouse antibodies to total cytokeratin (clone AE1/AE3, 'Dako'), HLA-DR (clone TAL.1B5, 'Dako'), and CD138 (clone MI15, 'Dako') were used as primary antibodies. RESULTS: The histological examination of the placentas in the IVF-S group showed the high incidence of central ischemic heart attacks (69%), dissociated cotyledon development (61%), pathological villous immaturity mainly with the predominance of intermediate differentiated villi (46%), and massive perivillous fibrinoid deposition (73%). The obtained differences between with the study and comparison groups were significant (p<0.05). The IVF-S group was characterized by the development of lymphoplasmacytic deciduitis (1.23±0.4 and 0.5±0.3 scores). Examination of the placental site biopsy material in the IVF-S group revealed the following changes: remodeling of the spiral arteries was incomplete in more than 40% of cases, and in 30% of the spiral arteries had no gestational changes. In the comparison group, more than 90% of the spiral arteries were characterized by complete remodeling during pregnancy. There was also an increase in the count of multinucleated trophoblastic giant cells (104.56±4.21 and 65.67±14.45) and HLA-DR positive cells (41.86±5.32 and 29.00±1.87). CONCLUSION: The placentas and the placental sites of the women whose pregnancy occurred with IVF-S are characterized by the development of high lyoplasmacytic deciduitis activity and pronounced placental immune alterations manifested by the high incidence of immune responses at the sites of the closest contact between maternal and fetal tissues. The placental bed exhibited defective spiral artery remodeling, development of chronic inflammatory lesions in the perivascular areas, and an increase in the counts of HLA-DR positive cells and multinucleated trophoblastic giant cells.
Subject(s)
Fertilization in Vitro , Placenta , Animals , Chorion , Female , Humans , Mice , Pregnancy , Tissue Donors , TrophoblastsABSTRACT
UNLABELLED: Renal cell carcinoma (RCC) is the most common malignant tumor of kidney associated with the worst clinical outcome. No molecular markers for RCC diagnostics and prognosis that could be applied in clinics were described yet. Large-scale screening of 3p human chromosome genes/loci in RCC and histologically normal tissues surrounding the tumors using NotI-microarray approach demonstrated that NKIRAS1 gene contained the largest percent of genetic/epigenetic changes in RCC tumor cells. AIM: To validate the results of NotI microarray analysis and study genetic, epigenetic changes, and the expression level of NKIRAS1 gene in human RCC samples. METHODS: DNA and RNA were isolated from freshly-frozen renal tumors' samples (n = 12) and from normal tissues surrounding the tumors. Epigenetic changes (methylation status) of NKIRAS1 were detected by bisulfite sequencing. Genetic changes and expression level were analyzed by Quantitative real-time PCR (qPCR) with SYBR Green. For relative quantification 2-(DeltaDeltaCP) method was used. Nonparametric tests (Wilcoxon, Kruskal - Wallis and Mann - Whitney) were applied for statistical data analysis using the BioStat software. RESULTS: NKIRAS1 expression was downregulated in 75% of RCC samples (9 of 12) compared with surrounding normal tissue. High grade tumors (3 and 4) showed lower expression of NKIRAS1 at the mRNA level than tumors of low grade (1 and 2). No significant association was found between gene expression level and gender or age. Analysis of NKIRAS1 gene copy number was performed in 19 tumor samples. Changes in the copy number of NKIRAS1 gene were observed in 64% (9 of 14) of cRCC samples. 9 samples displayed ratio (< 0.85 and >or= 0.35), thus were considered as hemizygous deletions. 3 samples showed ratio (> 0.85) and were considered as normal copy number. Changes in NKIRAS1 gene copy number were detected in all 3 benign oncocytomas, 1 papillary cancer and 1 sarcoma, where hemizygous deletion was observed. No changes in methylation status of NKIRAS1 were found in RCC. CONCLUSIONS: We have validated the results of NotI microarray analysis of NKIRAS1 gene in RCC. It was shown the decreased expression level of NKIRAS1 in this type of tumor.
