Assessment of the FAM174A 11G allele as a risk allele for equine metabolic syndrome.

An 11G nucleotide repeat in the 3' UTR of FAM174A was recently postulated as a risk allele with a dominant mode of inheritance for equine metabolic syndrome (EMS) and laminitis status in Arabian horses. The objective of this project was to evaluate this hypothesis in a large and diverse across-breed population. A total of 301 ponies, 292 Morgans, 64 Arabians, 49 Tennessee Walking Horses and 59 Quarter Horses were genotyped for six observed G repeat alleles in the FAM174A 3' UTR. Phenotype data included laminitis status, baseline insulin, glucose, non-esterified fatty acids, triglycerides, adiponectin, leptin, ACTH, insulin and glucose post oral sugar test, and two proxies for insulin resistance. The 11G allele frequencies were 18.8, 6.9, 1.8, 0.2 and 0.0% in the Arabians, Tennessee Walkers, ponies, Morgans and Quarter Horses respectively. Association analyses between FAM174A genotype and EMS phenotypes, and between allele count and EMS phenotypes, identified no statistically significant associations. When a dominant effect for the 11G allele was evaluated, a statistically significant association with adiponectin levels was identified in the ponies, and pairwise comparisons revealed that the estimated marginal means were higher in ponies with the 11G allele vs. alternative alleles (i.e. the allele had a protective effect). In conclusion, our data do not support the FAM174A 11G allele as a risk allele for EMS in our studied breeds.

EGFR mutation subtypes and response to immune checkpoint blockade treatment in non-small-cell lung cancer.

BACKGROUND: Although EGFR mutant tumors exhibit low response rates to immune checkpoint blockade overall, some EGFR mutant tumors do respond to these therapies; however, there is a lack of understanding of the characteristics of EGFR mutant lung tumors responsive to immune checkpoint blockade. PATIENTS AND METHODS: We retrospectively analyzed de-identified clinical and molecular data on 171 cases of EGFR mutant lung tumors treated with immune checkpoint inhibitors from the Yale Cancer Center, Memorial Sloan Kettering Cancer Center, University of California Los Angeles, and Dana Farber Cancer Institute. A separate cohort of 383 EGFR mutant lung cancer cases with sequencing data available from the Yale Cancer Center, Memorial Sloan Kettering Cancer Center, and The Cancer Genome Atlas was compiled to assess the relationship between tumor mutation burden and specific EGFR alterations. RESULTS: Compared with 212 EGFR wild-type lung cancers, outcomes with programmed cell death 1 or programmed death-ligand 1 (PD-(L)1) blockade were worse in patients with lung tumors harboring alterations in exon 19 of EGFR (EGFRΔ19) but similar for EGFRL858R lung tumors. EGFRT790M status and PD-L1 expression did not impact response or survival outcomes to immune checkpoint blockade. PD-L1 expression was similar across EGFR alleles. Lung tumors with EGFRΔ19 alterations harbored a lower tumor mutation burden compared with EGFRL858R lung tumors despite similar smoking history. CONCLUSIONS: EGFR mutant tumors have generally low response to immune checkpoint inhibitors, but outcomes vary by allele. Understanding the heterogeneity of EGFR mutant tumors may be informative for establishing the benefits and uses of PD-(L)1 therapies for patients with this disease.

Inclusion of herdmate data improves genomic prediction for milk-production and feed-efficiency traits within North American dairy herds.

Genomic data are widely available in the dairy industry and provide a cost-effective means of predicting genetic merit to inform selection decisions and increase genetic gains. As more dairy farms adopt genomic selection practices, dairy producers will soon have genomic data available on all of the animals within their herds. This is a very rich, but currently underused, source of information. Herdmates provide an excellent indication of how a selection candidate's genetics will perform within a given herd, noting that herdmates often include close relatives that share a similar environment. The study objective was to evaluate the utility of incorporating herdmate data into genomic predictions in a data set composed of 3,303 Holsteins from one herd in Canada and 6 herds throughout the United States. Within-herd prediction accuracy was assessed for milk-production and feed-efficiency traits determined from genomic best linear unbiased prediction under 4 different scenarios. Scenario 1 did not include herdmates in the training population. Scenarios 2 through 4 included herdmates in the training population, and scenarios 3 and 4 also included modeling of herd-specific marker effects. Leave-one-out cross validation was used to maximize the number of herdmates in the training population in scenarios 2 through 4, while maintaining constant training population size with scenario 1. Results from the present study reveal the importance of incorporating herdmate data into genomic evaluations. Inclusion of herdmates in the training population improved mean within-herd prediction accuracy for milk-production traits (± standard error) by 0.08 ± 0.03 (milk yield), 0.07 ± 0.03 (fat percentage), and 0.05 ± 0.01 (protein percentage) and feed-efficiency traits by 0.07 ± 0.02 (milk energy), 0.03 ± 0.02 (DMI), and 0.08 ± 0.01 (metabolic body weight). Modeling herd-specific marker effects further improved mean within-herd prediction accuracy for milk yield and energy by 0.03 ± 0.01 and 0.02 ± 0.01, respectively. Herds with higher within-herd heritability and low genomic correlation with the remaining herds benefitted most from the inclusion of herdmate data.

A framework to rank genomic alterations as targets for cancer precision medicine: the ESMO Scale for Clinical Actionability of molecular Targets (ESCAT).

Background: In order to facilitate implementation of precision medicine in clinical management of cancer, there is a need to harmonise and standardise the reporting and interpretation of clinically relevant genomics data. Methods: The European Society for Medical Oncology (ESMO) Translational Research and Precision Medicine Working Group (TR and PM WG) launched a collaborative project to propose a classification system for molecular aberrations based on the evidence available supporting their value as clinical targets. A group of experts from several institutions was assembled to review available evidence, reach a consensus on grading criteria and present a classification system. This was then reviewed, amended and finally approved by the ESMO TR and PM WG and the ESMO leadership. Results: This first version of the ESMO Scale of Clinical Actionability for molecular Targets (ESCAT) defines six levels of clinical evidence for molecular targets according to the implications for patient management: tier I, targets ready for implementation in routine clinical decisions; tier II, investigational targets that likely define a patient population that benefits from a targeted drug but additional data are needed; tier III, clinical benefit previously demonstrated in other tumour types or for similar molecular targets; tier IV, preclinical evidence of actionability; tier V, evidence supporting co-targeting approaches; and tier X, lack of evidence for actionability. Conclusions: The ESCAT defines clinical evidence-based criteria to prioritise genomic alterations as markers to select patients for targeted therapies. This classification system aims to offer a common language for all the relevant stakeholders in cancer medicine and drug development.

Clinical and molecular characterization of patients with cancer of unknown primary in the modern era.

BACKGROUND: On the basis of historical data, patients with cancer of unknown primary (CUP) are generally assumed to have a dismal prognosis with overall survival of less than 1 year. Treatment is typically cytotoxic chemotherapy guided by histologic features and the pattern of metastatic spread. The purpose of this study was to provide a clinical and pathologic description of patients with CUP in the modern era, to define the frequency of clinically actionable molecular alterations in this population, to determine how molecular testing can alter therapeutic decisions, and to investigate novel uses of next-generation sequencing in the evaluation and treatment of patients with CUP. PATIENTS AND METHODS: Under Institutional Review Board approval, we identified all CUP patients evaluated at our institution over a recent 2-year period. We documented demographic information, clinical outcomes, pathologic evaluations, next-generation sequencing of available tumor tissue, use of targeted therapies, and clinical trial enrollment. RESULTS: We identified 333 patients with a diagnosis of CUP evaluated at our institution from 1 January 2014 through 30 June 2016. Of these patients, 150 had targeted next-generation sequencing carried out on available tissue. Median overall survival in this cohort was 13 months. Forty-five of 150 (30%) patients had potentially targetable genomic alterations identified by tumor molecular profiling, and 15 of 150 (10%) received targeted therapies. Dominant mutation signatures were identified in 21 of 150 (14%), largely implicating exogenous mutagen exposures such as ultraviolet radiation and tobacco. CONCLUSIONS: Patients with CUP represent a heterogeneous population, harboring a variety of potentially targetable alterations. Next-generation sequencing may provide an opportunity for CUP patients to benefit from novel personalized therapies.

Oncologist use and perception of large panel next-generation tumor sequencing.

BACKGROUND: Genomic profiling is increasingly incorporated into oncology research and the clinical care of cancer patients. We sought to determine physician perception and use of enterprise-scale clinical sequencing at our center, including whether testing changed management and the reasoning behind this decision-making. PATIENTS AND METHODS: All physicians who consented patients to MSK-IMPACT, a next-generation hybridization capture assay, in tumor types where molecular profiling is not routinely performed were asked to complete a questionnaire for each patient. Physician determination of genomic 'actionability' was compared to an expertly curated knowledgebase of somatic variants. Reported management decisions were compared to chart review. RESULTS: Responses were received from 146 physicians pertaining to 1932 patients diagnosed with 1 of 49 cancer types. Physicians indicated that sequencing altered management in 21% (331/1593) of patients in need of a treatment change. Among those in whom treatment was not altered, physicians indicated the presence of an actionable alteration in 55% (805/1474), however, only 45% (362/805) of these cases had a genomic variant annotated as actionable by expert curators. Further evaluation of these patients revealed that 66% (291/443) had a variant in a gene associated with biologic but not clinical evidence of actionability or a variant of unknown significance in a gene with at least one known actionable alteration. Of the cases annotated as actionable by experts, physicians identified an actionable alteration in 81% (362/445). In total, 13% (245/1932) of patients were enrolled to a genomically matched trial. CONCLUSION: Although physician and expert assessment differed, clinicians demonstrate substantial awareness of the genes associated with potential actionability and report using this knowledge to inform management in one in five patients. CLINICAL TRIAL NUMBER: NCT01775072.

The performance of BRCA1 immunohistochemistry for detecting germline, somatic, and epigenetic BRCA1 loss in high-grade serous ovarian cancer.

BACKGROUND: BRCA1 expression can be lost by a variety of mechanisms including germline or somatic mutation and promotor hypermethylation. Given the potential importance of BRCA1 loss as a predictive and prognostic biomarker in high-grade serous ovarian cancer, we sought to evaluate the utility of BRCA1 immunohistochemistry (IHC) in screening for BRCA1 loss by germline, somatic, and epigenetic mechanisms. PATIENTS AND METHODS: Patients with advanced high-grade serous ovarian cancer who had previously undergone germline BRCA1 testing were identified. Samples from each tumor were stained for BRCA1 and reviewed independently by two pathologists blinded to BRCA status. Tumors with abnormal BRCA1 IHC and wild-type germline testing underwent further evaluation for somatic BRCA1 mutations and promoter hypermethylation. McNemar's test was used to determine the association of BRCA1 IHC with germline BRCA1 mutations and BRCA1 loss through any mechanism. Kaplan-Meier methods were used to estimate overall survival (OS), and the log-rank test was used to assess differences between groups. RESULTS: Inter-rater reliability between the two pathologists on BRCA IHC interpretation was very good (kappa coefficient 0.865, P = 0.16; McNemar's test). BRCA1 IHC was abnormal in 36% (48/135) of cases. When compared with germline BRCA1 status, BRCA1 IHC had a high negative predictive value (95.4%) but a low positive predictive value (PPV, 52.1%). When accounting for promoter hypermethylation and somatic mutations as alternative methods of BRCA1 loss, the PPV rose to 87.5%. Five-year OS rate was 49.6% [95% confidence interval (CI) 26.3% to 69.3%] for patients with germline BRCA1 mutations, 50.4% (95% CI 27.5% to 69.5%) for germline wild-type BRCA1 and abnormal IHC, and 52.1% (95% CI 38.4% to 64.2%) for germline wild-type BRCA1 and normal IHC (P = 0.92). CONCLUSIONS: BRCA1 IHC interpretation was a highly reproducible and accurate modality for detecting germline, somatic, or epigenetic mechanisms of BRCA1 loss. These results support further development of BRCA1 IHC as a potential biomarker for BRCA1 loss in high-grade serous ovarian cancer.

Evaluation of a fast-track programme for patients undergoing liver resection.

BACKGROUND: Recent developments in perioperative pathophysiology and care have documented evidence-based, multimodal rehabilitation (fast-track) to hasten recovery and to decrease morbidity and hospital stay for several major surgical procedures. The aim of this study was to investigate the effect of introducing fast-track principles for perioperative care in unselected patients undergoing open or laparoscopic liver resection. METHODS: This was a prospective study involving the first 100 consecutive patients who followed fast-track principles for liver resection. Catheters and drains were systematically removed early, and patients were mobilized and started eating and drinking from the day of surgery. An opioid-sparing multimodal pain treatment was given for the first week. Discharge criteria were: pain sufficiently controlled by oral analgesics alone, patient comfortable with discharge and no untreated complications. RESULTS: Median length of stay (LOS) for all patients was 5 days, with 2 days after laparoscopic versus 5 days following open resection (P < 0·001). Median LOS after minor open resections (fewer than 3 segments) was 5 days versus 6 days for major resections (3 or more segments) (P < 0·001). Simple right or left hemihepatectomies had a median LOS of 5 days. The readmission rate was 6·0 per cent and 30-day mortality was zero. CONCLUSION: Fast-track principles for perioperative care were introduced successfully and are safe after liver resection. Routine discharge 2 days after laparoscopic resection and 4-5 days after open liver resection may be feasible.

Assessing clinical researchers' information needs to create responsive portals and tools: my Research Assistant (MyRA) at the University of Utah: a case study.

QUESTION: How can health sciences librarians and biomedical informaticians offer relevant support to Clinical and Translational Science Award (CTSA) personnel? SETTING: The Spencer S. Eccles Health Sciences Library and the associate vice president for information technology for the health sciences office at the University of Utah conducted a needs assessment. METHODS: Faculty and staff from these two units, with the services of a consultant and other CTSA partners, employed a survey, focus groups, interviews, and committee discussions. An information portal was created to meet identified needs. RESULTS: A directive white paper was created. The process employed to plan a virtual and physical collaborative, collegial space for clinical researchers at the university and its three inter-institutional CTSA partners is described. CONCLUSION: The university's model can assist other librarians and informaticians with how to become part of a CTSA-focused infrastructure for clinical and translational research and serve researchers in general.

Preoperative high-dose glucocorticoids for early recovery after liver resection: randomized double-blinded trial.

BACKGROUND: Glucocorticoids modulate the surgical stress response. Previous studies showed that high-dose preoperative glucocorticoids reduce levels of postoperative inflammatory markers and specific biomarkers of liver damage compared with placebo, and suggested a reduced complication rate and shorter hospital stay after liver surgery. However, there are no studies with a clinical primary outcome or of early recovery outcomes. The aim of this study was to investigate whether a single high dose of preoperative glucocorticoid reduces complications in the immediate postoperative phase after liver surgery. METHODS: This was a single-centre, double-blinded, parallel-group RCT investigating preoperative methylprednisolone 10 mg/kg (high dose) versus dexamethasone 8 mg (standard-dose postoperative nausea prophylaxis) in patients scheduled for open liver resection. The primary outcome was number of patients with a complication in the postanaesthesia care unit; secondary outcomes included duration of hospital stay, pain and nausea during admission, and 30-day morbidity. RESULTS: A total of 174 patients (88 in high-dose group, 86 in standard-dose group) were randomized and analysed (mean(s.d.) age 65(12) years, 67.2 per cent men); 31.6 per cent had no serious co-morbidities and 25.3 per cent underwent major liver resection. Complications occurred in the postanaesthesia care unit in 51 patients (58 per cent) in the high-dose group and 58 (67 per cent) in the standard-dose group (risk ratio 0.86, 95 per cent c.i. 0.68 to 1.08; P = 0.213). Median duration of hospital stay was 4 days in both groups (P = 0.160). Thirty-day morbidity and mortality rates were similar in the two groups. CONCLUSION: A high dose of preoperative glucocorticoids did not reduce acute postoperative complications after open liver resection compared with a standard dose. Registration number: NCT03403517 (http://www.clinicaltrials.gov); EudraCT 2017-002652-81 (https://eudract.ema.europa.eu/).

MAPKs are essential upstream signaling pathways in proteolytic cartilage degradation--divergence in pathways leading to aggrecanase and MMP-mediated articular cartilage degradation.

OBJECTIVES: Matrix metalloproteinases (MMPs) and aggrecanases are essential players in cartilage degradation. However, the signaling pathways that results in MMP and/or aggrecanase synthesis and activation are not well understood. We investigated the molecular events leading to MMP- and aggrecanase-mediated cartilage degradation. METHODS: Cartilage degradation was induced in bovine articular cartilage explants by oncostatin M (OSM) and tumor necrosis factor (TNF), in the presence or absence of specific inhibitors of the mitogen-activated protein kinases (MAPKs) P38, P44/42 and Src family. Toxicity was followed by the AlamarBlue colorimetric assay. MMP-activity was assessed using a fluorescent substrate assay and MMP-9 and -2 activities by gelatinase zymography. MMP-mediated collagen type II degradation and MMP as well as aggrecanase-mediated aggrecan degradation was investigated with specific ELISA and hydroxyproline release by standard methods. The findings were verified by immunohistochemistry and histology. RESULTS: Stimulation of cartilage degradation by OSM+TNF resulted in 100-fold induction of CTX-II release (P<0.01). This was dose-dependently inhibited by MAPK P38 inhibitors and by the MAPK P44/42 inhibitors. MMP-activity and expression was significantly decreased, as evaluated by cleavage of fluorescence MMP-substrate and zymography. Immunohistochemistry confirmed these findings. Interestingly, only the P44/42 inhibitors abrogated aggrecanase-mediated aggrecan degradation. CONCLUSION: We found that inhibition of MAPK P38, P44/42 and Src family abrogated proteolytic cartilage degradation by blocking MMP synthesis and activity. However, only MAPK P44/42 was essential for aggrecanase-mediated aggrecan degradation. These data suggest that various aspects of cartilage degradation can be targeted independently by inhibiting specific upstream signaling pathway.

US oocyte donors: a retrospective study of medical and psychosocial issues.

BACKGROUND: First-person reports of oocyte donors, years after their donation, can give valuable information about medical complications of oocyte donation, as well as changes potentially required in procedures and priorities of US-based in vitro fertilization (IVF) centers. This paper reports findings from an online survey of former oocyte donors. METHODS: The instrument was an author-constructed questionnaire completed online on the Donor Sibling Registry website. Questions assessed women's accounts of medical complications, contact with the infertility clinic through which they had provided ova, and information exchange or contact with people conceived from their ova. RESULTS: Responses were received from 49.1% of the 287 donors with valid e-mail addresses. The 155 respondents completed the survey an average of 9.4 years after their first donation. Reported medical complications included ovarian hypersensitivity syndrome (30.3%) and infertility (9.6%). Subsequent to ova donation, 2.6% of women reported that they had been contacted by the IVF clinic for medical updates. On the questionnaire, 34.2% of women reported that medical changes they thought would interest donor children; half said that they had attempted to report these changes to the clinic with variable results. Many, who did not report such information, did not realize they could or should. Donors said that they frequently had not sought information about pregnancy outcomes because of confusion about the definition of 'anonymity' or 'confidentiality'. CONCLUSIONS: US-based IVF clinics need to give clearer guidelines to anonymous oocyte donors about follow-up information exchange. Additional long-term studies are needed to ascertain oocyte donors' risks of infertility or cancer.

Movement of a karyophilic protein through the nuclear pores of oocytes.

It has recently been shown that large karyophilic proteins are transported across the nuclear envelope in amphibian oocytes. In consideration of this, the present experiments were performed to identify the specific sites within the envelope through which transport occurs and determine if molecular size is a limiting factor in the transport process. The following experimental procedure was employed: Colloidal gold particles, varying in size from approximately 20 to 170 A in diameter were coated with nucleoplasmin, a 165,000-mol-wt karyophilic protein, which is known to be transported through the envelope. The coated gold particles were microinjected into the cytoplasm of Xenopus oocytes, and the cells were fixed 15 min and 1 h later. The intracellular localization of the gold was then determined with the electron microscope. It was found that nucleoplasmin-coated particles readily enter the nucleus. On the basis of the distribution of the particles associated with the envelope, we concluded that transport occurs through the nuclear pores. Furthermore, the size distributions of the gold particles present in the nucleus and cytoplasm were not significantly different, indicating that the envelope does not discriminate among particles with diameters ranging from 50 to 200 A (the dimensions including the nucleoplasmin coat). Colloidal gold coated with trypsin-digested nucleoplasmin (which lacks the polypeptide domain required for transport) or exogenous polyvinylpyrrolidone were largely excluded from the nucleus and showed no evidence of transport.

Heritability of metabolic traits associated with equine metabolic syndrome in Welsh ponies and Morgan horses.

BACKGROUND: Equine metabolic syndrome (EMS) is a complex clinical disorder with both environmental and genetic factors contributing to EMS phenotypes. Estimates of heritability determine the proportion of variation in a trait that is attributable to genetics. OBJECTIVES: To provide heritability estimates for nine metabolic traits associated with EMS in two high-risk breeds. STUDY DESIGN: Retrospective cohort study. METHODS: High-density single-nucleotide polymorphism (SNP) genotype data was used to estimate the heritability (h2 SNP ) of nine metabolic traits relevant to EMS in a cohort of 264 Welsh ponies and 286 Morgan horses. Traits included measurements of insulin, glucose, non-esterified fatty acids (NEFA), triglycerides, leptin, adiponectin, ACTH, and glucose (GLU-OST) and insulin (INS-OST) following an oral sugar challenge. RESULTS: In Welsh ponies, seven of the nine traits had statistically significant h2 SNP estimates that were considered moderately to highly heritable (h2 SNP >0.20) including: triglycerides (0.313; s.e. = 0.146), glucose (0.408; s.e. = 0.135), NEFA (0.434; s.e. = 0.136), INS-OST (0.440; s.e. = 0.148), adiponectin (0.488; s.e. = 0.143), leptin (0.554; s.e. = 0.132) and insulin (0.808; s.e. = 0.108). In Morgans, six of the nine traits had statistically significant h2 SNP estimates that were also determined to be moderately to highly heritable including: INS-OST (0.359; s.e. = 0.185), leptin (0.486; s.e. = 0.177), GLU-OST (0.566 s.e. = 0.175), insulin (0.592; s.e. = 0.195), NEFA (0.684; s.e. = 0.164), and adiponectin (0.913; s.e. = 0.181). MAIN LIMITATIONS: Insufficient population size may have limited power to obtain statistically significant h2 SNP estimates for ACTH (both breeds), glucose and triglycerides in Morgans and GLU-OST in Welsh ponies. CONCLUSIONS: This study provides the first concrete evidence of a genetic contribution to key phenotypes associated with EMS. Eight of these nine traits had moderate to high h2 SNP estimates in this cohort. These data demonstrate that continued research for identification of the genetic risk factors for EMS phenotypes within and across breeds is warranted.

Associations between endocrine disrupting chemicals and equine metabolic syndrome phenotypes.

Equine Metabolic Syndrome (EMS) is characterized by abnormalities in insulin regulation, increased adiposity and laminitis, and has several similarities to human metabolic syndrome. A large amount of environmental variability in the EMS phenotype is not explained by commonly measured factors (diet, exercise, and season), suggesting that other environmental factors play a role in EMS development. Endocrine disrupting chemicals (EDCs) are associated with metabolic syndrome and other endocrine abnormalities in humans. This led us to hypothesize that EDCs are detectable in horse plasma and play a role in the pathophysiology of EMS. EDCs acting through the aryl hydrocarbon and estrogen receptors, were measured in plasma of 301 horses from 32 farms. The median (range) TEQ (2,3,7,8-TCDD equivalent) and EEQ (17ß-estradiol equivalent) were 19.29 pg/g (0.59-536.36) and 10.50 pg/ml (4.35-15000.00), respectively. TEQ was negatively associated with plasma fat extracted and batch analyzed. EEQ was positively associated with pregnancy and batch analyzed, and negatively associated with being male and superfund score ≤100 miles of the farm. Of particular interest, serum glucose and insulin, glucose and insulin post oral sugar challenge, and leptin concentrations were associated with EEQ, and serum triglyceride concentration was associated with TEQ. Overall, we demonstrated that EDCs are present in the plasma of horses and may explain some of the environmental variability in measured EMS phenotypes. This is the first example of EDCs being associated with clinical disease phenotype components in domestic animals.

RAD51C (RAD51L2) is involved in maintaining centrosome number in mitosis.

The RAD51C (RAD51L2) protein is one out of five RAD51 paralogs and forms a complex that includes either XRCC2 or XRCC3. Both of these complexes may have important functions in homologous recombination (HR). Here, we confirm that the frequency of DNA double-strand break (DSB)-induced HR is reduced in the RAD51C deficient cell line CL-V4B, in agreement with a role for RAD51C in HR. We report that mitotic RAD51C deficient CL-V4B cells also have an increased number of centrosomes in mitosis resulting in aberrant mitotic spindles. These data suggest that the RAD51C protein is important in maintaining correct centrosome numbers and that the complexes including RAD51C and XRCC2 or XRCC3 may be of importance in maintaining correct centrosome numbers in mitosis. Increased centrosome numbers following a RAD51C defect indicates that this protein might be important in preventing aneuploidy, suggesting that it could be a potential tumour suppressor in mammals.

Kinetics and dose-response of residual 53BP1/gamma-H2AX foci: co-localization, relationship with DSB repair and clonogenic survival.

PURPOSE: Recent studies revealed that some foci produced by phosphorylated histone 2A family member X (gamma-H2AX) and tumor suppressor p53 binding protein 1 (53BP1) that co-localize with radiation-induced DNA double-strand breaks (DSB) remain in cells at relatively long times after irradiation and indicated a possible correlation between cellular radiosensitivity and residual foci. In this study, we investigated dose-responses and kinetics for radiation-induced 53BP1/gamma-H2AX foci formation in relation to their co-localization, DSB repair and cell survival. MATERIALS AND METHODS: Cell survival, DSB and foci were analyzed by clonogenic assay, pulsed field gel electrophoresis (PFGE), and confocal laser microscopy, respectively, in normal human fibroblasts (VH-10) and in a cancer cell line (HeLa). Computer analysis was used to determine both the number and the area of foci. RESULTS: We show that even at doses down to 1 cGy a statistically significant induction of 53BP1 foci is observed. While the number of foci was found to constantly decrease with post-irradiation time, the per-cell normalized area of foci does not change within a time window of approximately 4 h post-irradiation. Co-localization of gamma-H2AX and 53BP1 foci is shown to depend on dose and post-irradiation time. No clear correlations were established between radiosensitivity and foci formation because the dose response for 53BP1/gamma-H2AX foci may depend on time after irradiation and duration of the cell cycle. We show that the kinetics of foci disappearance within 24 h post-irradiation do not coincide with those of DSB repair. CONCLUSIONS: The data suggest that the post-irradiation time used for estimation of radiosensitivity at therapeutically relevant low doses (e.g., <3 Gy) in proliferating cells by scoring residual foci should be limited by the duration of the cell cycle, and that direct comparison of the kinetics of DSB repair and disappearance of DSB-co-localizing foci is not possible. Therefore, results obtained from the counting of foci should be interpreted with caution in terms of DSB repair.

Posterior hypothalamic lesions advance the time of the pubertal changes in luteinizing hormone release in ovariectomized female rhesus monkeys.

To examine further the relationship between developmental changes in LH release and the onset of puberty, effects of posterior hypothalamic lesions were tested in ovariectomized (OVX), sexually immature female monkeys. In OVX females (n = 3) with sham hypothalamic lesions basal LH levels were suppressed during the prepubertal period until 25 months of age, when LH levels started to increase. The increase in basal LH continued; a 100% elevation from prepubertal levels was attained at 26.0 +/- 0 months of age, and a 200% elevation was attained at 31.0 +/- 3.2 months of age. A consistent appearance of LH circadian fluctuation (nocturnal LH increase) with a large amplitude accompanied the initial LH increase. Lesions of the posterior hypothalamus (PH) in OVX animals (n = 6) at 17-18 months of age, which we previously reported to be effective in advancing the onset of puberty by several months in ovarian intact monkeys, resulted in an early 100% increase in basal LH levels and the circadian LH fluctuation (19.5 +/- 1.0 months of age). Basal LH levels in these animals further increased, reaching a 200% elevation of prelesion levels at 24.2 +/- 0.7 months of age. All of these LH changes with PH lesions occurred significantly (P less than 0.01) earlier than those in sham-lesioned animals. Lesion of the PH in OVX animals (n = 4) at 13-14 months of age resulted in an increase in LH and the circadian LH fluctuation within 1 month postoperatively. However, 100% and 200% LH elevations did not occur until 20.8 +/- 1.0 and 24.8 +/- 1.4 months of age, respectively. These ages were similar to those of animals receiving lesions at 17-18 months of age, but much younger than those of sham controls (P less than 0.01). PH lesions in animals at 13-14 months of age also advanced the time of the first positive feedback effects of estrogen. In animals (n = 4) with PH lesions, estradiol benzoate induced a first LH response at 21.5 +/- 1.6 months of age, when basal LH was 276 +/- 83% increased from prelesion levels. This age was significantly (P less than 0.05) younger than that (29.3 +/- 1.9 months; n = 6) of the first LH surge induced by estrogen in control animals when basal LH levels attained 248 +/- 18% of prepubertal levels.(ABSTRACT TRUNCATED AT 400 WORDS)

Norepinephrine is a possible neurotransmitter stimulating pulsatile release of luteinizing hormone-releasing hormone in the rhesus monkey.

The hypothesis that norepinephrine (NE) plays a facilitatory role in controlling the pulsatile release of LHRH was tested with a modified push-pull perfusion technique in conscious rhesus monkeys. The in vivo LHRH release in perfusate samples collected from the stalk-median eminence of ovariectomized females was pulsatile and synchronous with pulsatile LH release. Catecholamines measured in aliquots of perfusate samples revealed that in vivo NE release was also pulsatile and was synchronous with LHRH release. Local infusion of NE or methoxamine (an alpha 1-adrenergic stimulant) through a push cannula stimulated LHRH release, while iv injection of prazosin (an alpha 1-adrenergic blocker) suppressed LHRH release. It is concluded that NE is a possible neurotransmitter stimulating pulsatile LHRH release.

Clinical significance of cognitive performance by hypertensive patients.

Fifty-four subjects with uncomplicated essential hypertension and 54 normotensive subjects were compared with regard to a widely employed clinical index of cognitive dysfunction (the Average Impairment Rating) calculated from neuropsychological tests that discriminate between brain-damaged and neurologically normal persons. Hypertensive subjects exhibited lower mean scores on this index when education was ignored, but results were not the same for highly educated and less well educated groups. There were no differences between exceptionally well educated hypertensive and normotensive subjects, but in the less well educated group, hypertensive subjects performed more poorly than normotensive subjects. The percentages of hypertensive and normotensive subjects scoring in a cognitively impaired range on the Average Impairment Rating were low and did not differ for either education group. These data indicate the important role of subtle differences in education level with respect to positive or negative findings for studies comparing hypertensive and normotensive subjects and illustrate the important role of clinical neuropsychological indices of cognitive dysfunction when one wishes to make meaningful inferences regarding cerebral cortical function in hypertensive subjects.