Factors associated with aberrant imprint methylation and oligozoospermia.

Disturbingly, the number of patients with oligozoospermia (low sperm count) has been gradually increasing in industrialized countries. Epigenetic alterations are believed to be involved in this condition. Recent studies have clarified that intrinsic and extrinsic factors can induce epigenetic transgenerational phenotypes through apparent reprogramming of the male germ line. Here we examined DNA methylation levels of 22 human imprinted loci in a total of 221 purified sperm samples from infertile couples and found methylation alterations in 24.8% of the patients. Structural equation model suggested that the cause of imprint methylation errors in sperm might have been environmental factors. More specifically, aberrant methylation and a particular lifestyle (current smoking, excess consumption of carbonated drinks) were associated with severe oligozoospermia, while aging probably affected this pathology indirectly through the accumulation of PCB in the patients. Next we examined the pregnancy outcomes for patients when the sperm had abnormal imprint methylation. The live-birth rate decreased and the miscarriage rate increased with the methylation errors. Our research will be useful for the prevention of methylation errors in sperm from infertile men, and sperm with normal imprint methylation might increase the safety of assisted reproduction technology (ART) by reducing methylation-induced diseases of children conceived via ART.

Epigenetic alterations in sperm associated with male infertility.

The most common form of male infertility is a low sperm count, known as oligozoospermia. Studies suggest that oligozoospermia is associated with epigenetic alterations. Epigenetic alterations in sperm, which may arise due to the exposure of gametes to environmental factors or those that pre-exist in the sperm of infertile individuals, may contribute to the increased incidence of normally rare imprinting disorders in babies conceived after assisted reproductive technology using the sperm of infertile men. Genomic imprinting is an important developmental process whereby the allelic activity of certain genes is regulated by DNA methylation established during gametogenesis. The aberrant expression of several imprinted genes has been linked to various diseases, malignant tumors, lifestyle and mental disorders in humans. Understanding how infertility and environmental factors such as reproductive toxicants, certain foods, and drug exposures during gametogenesis contribute to the origins of these disorders via defects in sperm is of paramount importance. In this review, we discuss the association of epigenetic alterations with abnormal spermatogenesis and the evidence that epigenetic processes, including those required for genomic imprinting, may be sensitive to environmental exposures during gametogenesis, fertilization and early embryonic development. In addition, we review imprinting diseases and their relationships with environmental factors. While the plasticity of epigenetic marks may make these more susceptible to modification by the environment, this also suggests that aberrant epigenetic marks may be reversible. A greater understanding of this process and the function of epidrugs may lead to the development of new treatment methods for many adult diseases in the future.

Stability of genomic imprinting in human induced pluripotent stem cells.

BACKGROUND: hiPSCs are generated through epigenetic reprogramming of somatic tissue. Genomic imprinting is an epigenetic phenomenon through which monoallelic gene expression is regulated in a parent-of-origin-specific manner. Reprogramming relies on the successful erasure of marks of differentiation while maintaining those required for genomic imprinting. Loss of imprinting (LOI), which occurs in many types of malignant tumors, would hinder the clinical application of hiPSCs. RESULTS: We examined the imprinting status, expression levels and DNA methylation status of eight imprinted genes in five independently generated hiPSCs. We found a low frequency of LOI in some lines. Where LOI was identified in an early passage cell line, we found that this was maintained through subsequent passages of the cells. Just as normal imprints are maintained in long-term culture, this work suggests that abnormal imprints are also stable in culture. CONCLUSIONS: Analysis of genomic imprints in hiPSCs is a necessary safety step in regenerative medicine, with relevance both to the differentiation potential of these stem cells and also their potential tumorigenic properties.

High-throughput detection of aberrant imprint methylation in the ovarian cancer by the bisulphite PCR-Luminex method.

BACKGROUND: Aberrant DNA methylation leads to loss of heterozygosity (LOH) or loss of imprinting (LOI) as the first hit during human carcinogenesis. Recently we developed a new high-throughput, high-resolution DNA methylation analysis method, bisulphite PCR-Luminex (BPL), using sperm DNA and demonstrated the effectiveness of this novel approach in rapidly identifying methylation errors. RESULTS: In the current study, we applied the BPL method to the analysis of DNA methylation for identification of prognostic panels of DNA methylation cancer biomarkers of imprinted genes. We found that the BPL method precisely quantified the methylation status of specific DNA regions in somatic cells. We found a higher frequency of LOI than LOH. LOI at IGF2, PEG1 and H19 were frequent alterations, with a tendency to show a more hypermethylated state. We detected changes in DNA methylation as an early event in ovarian cancer. The degree of LOI (LOH) was associated with altered DNA methylation at IGF2/H19 and PEG1. CONCLUSIONS: The relative ease of BPL method provides a practical method for use within a clinical setting. We suggest that DNA methylation of H19 and PEG1 differentially methylated regions (DMRs) may provide novel biomarkers useful for screening, diagnosis and, potentially, for improving the clinical management of women with human ovarian cancer.

[Genomic imprinting and carcinogenesis].

Genomic imprinting is an epigenetic marking and a stable transmission of monoallelic gene expression patterns in a parent of- origin-specific manner. Aberrant imprinting has been linked to a number of human genetic disorders, including congenital abnormalities, childhood cancer, behavior disorders, and cancer in adults. Imprinted genes play roles in carcinogenesis. Recently, progress in researched on epigenetic mechanisms of imprinted genes, in edition to analysis of the pathology of the oncogenetic mechanisms, has begun to be clinically applied to diagnostic methods, prevention, and cancer drug development.

High incidence of renal failure in patients with aortic aneurysms.

BACKGROUND: Renal failure (RF) is a well-recognized complication of aortic aneurysms (AA) although its incidence has been poorly documented previously. The purpose of this study is to examine the incidence of RF in patients with AA and prognosis of AA patients with RF. METHODS: Renal function, complications and prognosis of AA patients with RF were retrospectively reviewed in 350 AA patients (median age 69.8+/-10.7 years) in the International Medical Center of Japan from 1989 to 1999. RESULTS: Among 350 patients with AA, 90 patients (25.7%) had chronic renal failure (CRF) at the initiation of follow-up. The number of CRF patients increased to 117 (33.4%) at 30 months of follow-up. Forty-four out of 160 patients (27.5%) who had aortic surgery developed postoperative acute renal failure (ARF). Stepwise logistic regression analysis revealed that age (>or=65 years), hypertension and multiple aneurysms were independent risk factors for CRF, whereas dissecting aneurysms, preoperative serum creatinine (sCr) levels and duration of surgery were independent risk factors for postoperative ARF in AA patients. In the 5-year follow-up of AA patients with CRF, the mean slopes of 1/serum-creatinine did not significantly differ between conservative treatment and surgical treatment. The survival rates were 49.5% in the conservative treatment group and 67.3% in the surgical treatment group. CONCLUSION: Our data suggest that the management of renal function including blood pressure from an early stage in AA patients is important since CRF is highly prevalent in AA patients and affects their prognosis and mortality.

Synaptic vesicle protein 2B is expressed in podocyte, and its expression is altered in proteinuric glomeruli.

Synaptic vesicle protein 2B (SV2B) was identified by the subtraction hybridization technique as a molecule of which mRNA expression was decreased in puromycin aminonucleoside (PAN) nephropathy by glomerular cDNA subtraction assay. The expression of SV2B was detected in glomerular lysate with Western blot analysis. Dual-labeling immunofluorescence studies with glomerular cell markers demonstrated that SV2B is expressed in glomerular visceral epithelial cells (podocytes). The expression of SV2B is detected also in cultured podocyte and in human kidney section as podocytic pattern. The decrease of SV2B mRNA was already detected before the onset of proteinuria in PAN nephropathy. The mRNA expression of SV2B clearly is altered not only in PAN nephropathy but also in another proteinuric state that is caused by an antibody against nephrin, a functional molecule of the slit diaphragm. The decreased intensity in SV2B staining was already detected before the peak of proteinuria in both models with immunofluorescence study. A reduced amount of SV2B was detected in both models also with Western blot analysis. CD2AP, another functional molecule of the slit diaphragm, was observed in cytoplasm, including the processes area of the cultured podocyte, and when the podocyte was treated with small interfering RNA for SV2B, CD2AP staining at the process area was not detected. These results suggest that SV2B is a functional molecule of podocyte, and SV2B may play a role in the expression and proper localization of CD2AP.

ANCA-related crescentic glomerulonephritis in a patient with scleroderma without marked dermatological change and malignant hypertension.

A 64-year-old woman with scleroderma without marked dermatological change developed anti-neutrophil cytoplasmic autoantibody (ANCA)-related renal failure. She had neither malignant hypertension nor elevation of plasma renin concentration. Renal biopsy showed crescentic glomerulonephritis (pauci-immune type) and the myeloperoxidase-specific anti-neutrophil cytoplasmic autoantibody (MPO-ANCA) titer was found to be elevated to 757 IU/ml. Methylprednisolone pulse therapy followed by oral prednisolone effectively suppressed renal failure and lowered the MPO-ANCA titer. We believe this is a rare case of ANCA-related renal failure in a patient with scleroderma without marked dermatological change.