Heparin and penicillamine–hypotaurine–epinephrine (PHE) solution during bovine in vitro fertilization procedures impair the quality of spermatozoa but improve normal oocyte fecundation and early embryonic development

The presence of heparin and a mixture of penicillamine, hypotaurine, and epinephrine (PHE) solution in the in vitro fertilization (IVF) media seem to be a prerequisite when bovine spermatozoa are capacitated in vitro, in order to stimulate sperm motility and acrosome reaction. The present study was designed to determine the effect of the addition of heparin and PHE during IVF on the quality and penetrability of spermatozoa into bovine oocytes and on subsequent embryo development. Sperm quality, evaluated by the integrity of plasma and acrosomal membranes and mitochondrial function, was diminished (P<0.05) in the presence of heparin and PHE. Oocyte penetration and normal pronuclear formation rates, as well as the percentage of zygotes presenting more than two pronuclei, was higher (P<0.05) in the presence of heparin and PHE. No differences were observed in cleavage rates between treatment and control (P>0.05). However, the developmental rate to the blastocyst stage was increased in the presence of heparin and PHE (P>0.05). The quality of embryos that reached the blastocyst stage was evaluated by counting the inner cell mass (ICM) and trophectoderm (TE) cell numbers and total number of cells; the percentage of ICM and TE cells was unaffected (P>0.05) in the presence of heparin and PHE (P<0.05). In conclusion, this study demonstrated that while the supplementation of IVF media with heparin and PHE solution impairs spermatozoa quality, it plays an important role in sperm capacitation, improving pronuclear formation, and early embryonic development

Psychophysical analysis of contrast processing segregated into magnocellular and parvocellular systems in asymptomatic carriers of 11778 Leber`s hereditary optic neuropathy

We examined achromatic contrast discrimination in asymptomatic carriers of 11778 Leber`s hereditary optic neuropathy (LHON 18 controls) and 18 age-match were also tested. To evaluate magnocellular (MC) and Parvocellular (PC) contrast discrimination, we used a version of Pokorny and Smith`s (1997) Pulsed/steady-pedestal paradigms (PPP/SPP) thought to be detected via PC and MC pathways, respectively. A luminance pedestal (four 1 degrees x 1 degrees squares) was presented on a 12 cd/m(2) surround. The luminance of one of the squares (trial square, TS) was randomly incremented for either 17 or 133 ms. Observers had to detect the TS, in a forced-choice task, at each duration, for three pedestal levels: 7, 12, 19 cd/m(2). In the SPP, the pedestal was fixed, and the TS was modulated. For the PPP, all four pedestal squares pulsed for 17 or 133 ms, and the TS was simultaneously incremented or decremented. We found that contrast discrimination thresholds of LHON carriers were significantly higher than controls` in the condition with the highest luminance of both paradigms, implying impaired contrast processing with no evidence of differential sensitivity losses between the two systems. Carriers` thresholds manifested significantly longer temporal integration than controls in the SPP, consistent with slowed MC responses. The SPP and PPP paradigms can identify contrast and temporal processing deficits in asymptomatic LHON carriers, and thus provide an additional tool for early detection and characterization of the disease.

Prevalence of the BRCA1 founder mutation c.5266dupin Brazilian individuals at-risk for the hereditary breast and ovarian cancer syndrome

About 5-10% of breast and ovarian carcinomas are hereditary and most of these result from germline mutations in the BRCA1 and BRCA2 genes. In women of Ashkenazi Jewish ascendance, up to 30% of breast and ovarian carcinomas may be attributable to mutations in these genes, where 3 founder mutations, c.68_69del (185delAG) and c.5266dup (5382insC) in BRCA1 and c.5946del (6174delT) in BRCA2, are commonly encountered. It has been suggested by some authors that screening for founder mutations should be undertaken in all Brazilian women with breast cancer. Thus, the goal of this study was to determine the prevalence of three founder mutations, commonly identified in Ashkenazi individuals in a sample of non-Ashkenazi cancer-affected Brazilian women with clearly defined risk factors for hereditary breast and ovarian cancer (HBOC) syndrome. Among 137 unrelated Brazilian women from HBOC families, the BRCA1c.5266dup mutation was identified in seven individuals (5%). This prevalence is similar to that encountered in non-Ashkenazi HBOC families in other populations. However, among patients with bilateral breast cancer, the frequency of c.5266dup was significantly higher when compared to patients with unilateral breast tumors (12.1% vs 1.2%, p = 0.023). The BRCA1 c.68_69del and BRCA2 c.5946del mutations did not occur in this sample. We conclude that screening non-Ashkenazi breast cancer-affected women from the ethnically heterogeneous Brazilian populations for the BRCA1 c.68_69del and BRCA2 c.5946del is not justified, and that screening for BRCA1c.5266dup should be considered in high risk patients, given its prevalence as a single mutation. In high-risk patients, a negative screening result should always be followed by comprehensive BRCA gene testing. The finding of a significantly higher frequency of BRCA1 c.5266dup in women with bilateral breast cancer, as well as existence of other as yet unidentified founder mutations in this population, should be further assessed in a larger well characterized high-risk cohort.

Toward a Clinical Protocol for Assessing Rod, Cone, and Melanopsin Contributions to the Human Pupil Response

PURPOSE. To better understand the relative contributions of rod, cone, and melanopsin to the human pupillary light reflex (PLR) and to determine the optimal conditions for assessing the health of the rod, cone, and melanopsin pathways with a relatively brief clinical protocol. METHODS. PLR was measured with an eye tracker, and stimuli were controlled with a Ganzfeld system. In experiment 1, 2.5 log cd/m(2) red (640 +/- 10 nm) and blue (467 +/- 17 nm) stimuli of various durations were presented after dark adaptation. In experiments 2 and 3, 1-second red and blue stimuli were presented at different intensity levels in the dark (experiment 2) or on a 0.78 log cd/m(2) blue background (experiment 3). Based on the results of experiments 1 to 3, a clinical protocol was designed and tested on healthy control subjects and patients with retinitis pigmentosa and Leber`s congenital amaurosis. RESULTS. The duration for producing the optimal melanopsin-driven sustained pupil response after termination of an intense blue stimulus was 1 second. PLR rod-and melanopsin-driven components are best studied with low-and high-intensity flashes, respectively, presented in the dark (experiment 2). A blue background suppressed rod and melanopsin responses, making it easy to assess the cone contribution with a red flash (experiment 3). With the clinical protocol, robust melanopsin responses could be seen in patients with few or no contributions from the rods and cones. CONCLUSIONS. It is possible to assess the rod, cone, and melanopsin contributions to the PLR with blue flashes at two or three intensity levels in the dark and one red flash on a blue background. (Invest Ophthalmol Vis Sci. 2011; 52: 6624-6635) DOI: 10.1167/iovs.11-7586

Germline DNA copy number variation in familial and early-onset breast cancer

Introduction: Genetic factors predisposing individuals to cancer remain elusive in the majority of patients with a familial or clinical history suggestive of hereditary breast cancer. Germline DNA copy number variation (CNV) has recently been implicated in predisposition to cancers such as neuroblastomas as well as prostate and colorectal cancer. We evaluated the role of germline CNVs in breast cancer susceptibility, in particular those with low population frequencies (rare CNVs), which are more likely to cause disease." Methods: Using whole-genome comparative genomic hybridization on microarrays, we screened a cohort of women fulfilling criteria for hereditary breast cancer who did not carry BRCA1/BRCA2 mutations. Results: The median numbers of total and rare CNVs per genome were not different between controls and patients. A total of 26 rare germline CNVs were identified in 68 cancer patients, however, a proportion that was significantly different (P = 0.0311) from the control group (23 rare CNVs in 100 individuals). Several of the genes affected by CNV in patients and controls had already been implicated in cancer. Conclusions: This study is the first to explore the contribution of germline CNVs to BRCA1/2-negative familial and early-onset breast cancer. The data suggest that rare CNVs may contribute to cancer predisposition in this small cohort of patients, and this trend needs to be confirmed in larger population samples.

Hereditary Autoinflammatory Syndromes: A Brazilian Multicenter Study

To evaluate the prevalence of genetic defects in clinically suspected autoinflammatory syndromes (AIS) in a Brazilian multicenter study. The study included 102 patients with a clinical diagnosis of Cryopyrin Associated Periodic Syndromes (CAPS), TNF Receptor Associated Periodic Syndrome (TRAPS), Familial Mediterranean Fever (FMF), Mevalonate Kinase Deficiency (MKD) and Pediatric Granulomatous Arthritis (PGA). One of the five AIS-related genes (NLRP3, TNFRSF1A, MEFV, MVK and NOD2) was evaluated in each patient by direct DNA sequencing, based on the most probable clinical suspect. Clinical diagnoses of the 102 patients were: CAPS (n = 28), TRAPS (n = 31), FMF (n = 17), MKD (n = 17) and PGA (n = 9). Of them, 27/102 (26 %) had a confirmed genetic diagnosis: 6/28 (21 %) CAPS patients, 7/31 (23 %) TRAPS, 3/17 (18 %) FMF, 3/17 (18 %) MKD and 8/9 (89 %) PGA. We have found that approximately one third of the Brazilian patients with a clinical suspicion of AIS have a confirmed genetic diagnosis.

Mutation Spectrum in the Large GTPase Dynamin 2, and Genotype-Phenotype Correlation in Autosomal Dominant Centronuclear Myopathy

Centronuclear myopathy (CNM) is a genetically heterogeneous disorder associated with general skeletal muscle weakness, type I fiber predominance and atrophy, and abnormally centralized nuclei. Autosomal dominant CNM is due to mutations in the large GTPase dynamin 2 (DNM2), a mechanochemical enzyme regulating cytoskeleton and membrane trafficking in cells. To date, 40 families with CNM-related DNM2 mutations have been described, and here we report 60 additional families encompassing a broad genotypic and phenotypic spectrum. In total, 18 different mutations are reported in 100 families and our cohort harbors nine known and four new mutations, including the first splice-site mutation. Genotype-phenotype correlation hypotheses are drawn from the published and new data, and allow an efficient screening strategy for molecular diagnosis. In addition to CNM, dissimilar DNM2 mutations are associated with Charcot-Marie-Tooth (CMT) peripheral neuropathy (CMTD1B and CMT2M), suggesting a tissue-specific impact of the mutations. In this study, we discuss the possible clinical overlap of CNM and CMT, and the biological significance of the respective mutations based on the known functions of dynamin 2 and its protein structure. Defects in membrane trafficking due to DNM2 mutations potentially represent a common pathological mechanism in CNM and CMT. Hum Mutat 33: 949-959, 2012. (C) 2012 Wiley Periodicals, Inc.