Identification of a compound heterozygous missense mutation in LAMA2 gene from a patient with merosin‐deficient congenital muscular dystrophy type 1A

BackgroundMerosin‐deficient congenital muscular dystrophy type 1A (MDC1A) is occurred by mutations in LAMA2 gene that encodes the laminin α2 chain (merosin). MDC1A is a predominant subtype of congenital muscular dystrophy. Herein, we identified two missense mutations in LAMA2 gene in compound heterozygous status in an Iranian patient with MDC1A using whole‐exome sequencing (WES).MethodsIn the present study, we evaluated genetic alterations in an Iranian 35‐month‐old boy with MDC1A and his healthy family using WES method. The identified mutations further confirmed by Sanger sequencing method. Finally, in silico analysis was conducted to further evaluation of molecular function of the identified genetic variants.ResultsWe identified two potentially pathogenic missense mutations in compound heterozygous state (c.7681G>A p.Gly2561Ser and c.4840A>G p.Asn1614Asp) in LAMA2 gene as contributing to the MDC1A phenotype. The healthy parents of our proband are single heterozygous for identified mutations. These variants were found to be pathogenic by in silico analysis.ConclusionsIn general, we successfully identified LAMA2 gene mutations in an Iranian patient with MDC1A using WES. The identified mutations in LAMA2 gene can be useful in genetic counseling, prenatal diagnosis, and predicting prognosis of MDC1A.     

Effects of intraoperative steroid injection on the outcome of pterygium surgery

Purpose

To evaluate the effects of intraoperative triamcinolone injection on the outcome of pterygium surgery.

Methods

This prospective study included 54 eyes with primary nasal pterygia that underwent pterygium surgery with a bare-sclera technique and intraoperative mitomycin C application. Patients were randomized into two groups; the steroid group that received subconjunctival injection of 12 mg triamcinolone acetonide at the end of surgery, and the control group that did not receive such steroid injection. Main outcome measures included presence of conjunctival inflammation at 1 month postoperatively as well as recurrence of pterygium.

Results

Twelve-month follow-up was completed in 48 eyes (23 in the steroid group and 25 in the control group). At 1 month postoperatively, different grades of conjunctival inflammation were present in 11 (47.8%) of the steroid group and in 14 (56%) of the control group (P=0.39). For eyes with moderate or severe postoperative inflammation, subconjunctival triamcinolone was injected; these included 6 (26.1%) and 9 (36%) in the steroid and control groups, respectively (P=0.54). During follow-up, surgical area showed fine episcleral vessels without fibrous tissue in 1 (4.3%) of the steroid group and 3 (12.0%) of the control group (P=0.33), which all regressed after triamcinolone injection. Conjunctival recurrence of pterygium was seen in 2 (8.7%) of the steroid group and in 1 (4.0%) of the control group (P=0.47). No eye developed corneal recurrence in either group.

Conclusions

In pterygium surgery with a bare-sclera technique and mitomycin C application, intraoperative triamcinolone injection did not significantly reduce postoperative conjunctival inflammation or pterygium recurrence.     

Prophylactic corticosteroid use prevents engraftment syndrome in patients after autologous stem cell transplantation

Engraftment syndrome (ES) following autologous stem cell transplantation (ASCT) at the time of neutrophil recovery may comprise fever, rash, pulmonary edema, or diarrhea. Usually, ES is easily manageable using corticosteroids but may prolong hospitalization. In two consecutive cohorts of subsequent patients with myeloma, lymphomas, and testicular/germ cell cancer, we assessed the benefit of corticosteroid use to prevent incidence and severity of ES following ASCT. Whereas Cohort A (82 patients) received no prophylactic corticosteroids, corticosteroids (4 mg dexamethasone oral daily) were started in Cohort B (60 patients) at day +9 until day +13 following ASCT. Steroid prophylaxis significantly reduced the incidence of ES (6/60; 10% vs. 33/82; 40%; p < 0.001). Hospitalization duration was longer in patients with ES than in patients without ES within both cohorts (in Cohort A: p = 0.007; and B: p = 0.011), but did not differ significantly between cohorts A and B. Finally, in Cohort A, there was a trend to an inferior 2‐year overall survival rate in patients without ES compared to patients with ES (p = 0.067), but definite conclusions are not yet allowed. Our results suggest that corticosteroid prophylaxis from days +9 to +13 following ASCT significantly reduces the risk of ES and shortens hospitalization duration.     

Induction of DNA polymerase beta-dependent base excision repair in response to oxidative stress in vivo

Base excision repair (BER) is the DNA repair pathway primarily responsible for repairing small base modifications and abasic sites caused by normal cellular metabolism or environmental insult. Strong evidence supports the requirement of DNA polymerase beta (beta-pol) in the BER pathway involving single nucleotide gap filling DNA synthesis in mammalian systems. In this study, we examine the relationship between oxidative stress, cellular levels of beta-pol and BER to determine whether oxidizing agents can upregulate BER capacity in vivo. Intraperitoneal injection of 2-nitropropane (2-NP, 100 mg/kg), an oxidative stress-inducing agent, in C57BL/6 mice was found to generate 8-hydroxydeoxyguanosine (8-OHdG) in liver tissue (4-fold increase, P < 0.001). We also observed a 4-5-fold increase in levels of DNA single strand breaks in 2-NP treated animals. The protein level of the tumor suppressor gene, p53 was also induced in liver by 2-NP (2.1-fold, P < 0.01), indicating an induction of DNA damage. In addition, we observed a 2-3-fold increase in mutant frequency in the lacI gene after exposure to 2-NP. Interestingly, an increase in DNA damage upregulated the level of beta-pol as well as BER capacity (42%, P < 0.05). These results suggest that beta-pol and BER can be upregulated in response to oxidative stress in vivo. Furthermore, data show that heterozygous beta-pol knockout (beta-pol(+/-)) mice express higher levels of p53 in response to 2-NP as compared with wild-type littermates. While the knockout and wild-type mice display similar levels of 8-OHdG after 2-NP exposure, the beta-pol(+/-) mice exhibit a significant increase in DNA single strand breaks. These findings suggest that in mice, a reduction in beta-pol expression results in a higher accumulation of DNA damage by 2-NP, thus establishing the importance of the beta-pol-dependent BER pathway in repairing oxidative damage.     

Association of TNF-α and TNF-β gene polymorphism with steroid receptor expression in breast cancer patients

The presence of estrogen and progesterone receptors is correlated with good prognosis in breast cancer. The effect of TNF-α on down-regulation of estrogen receptor and blocking the proliferative response of breast cancer cells to estradiol have been demonstrated. However, the effect of TNFA and TNFB gene polymorphisms on the expression of steroid receptors in breast cancer cells is not well documented. Therefore, 160 breast cancer patients were recruited to investigate the association of TNFA and TNFB gene polymorphism with the level of steroid receptor expression. This association was not found to be significant for TNFA polymorphism and estrogen receptor expression (p=0.07). However, when combined genotypes of TNFA and TNFB polymorphism was considered, homozygous patients for lower TNF-α producer genotypes (TNFA1/A1 and TNFB1/B1) showed significantly higher progesterone receptor expression (p=0.041). Our findings indicate that TNFA and TNFB polymorphisms may be associated with the levels of steroid receptor expression in breast cancer patients. Further studies on a larger group of breast cancer patients are recommended.     

Evaluation of intraocular penetration of levofloxacin by high performance liquid chromatography

AIM: To evaluate the levofloxacin eye drop into human eye penetration, levofloxacin eye drop concentrations in human ocular aqueous of 33 patients undergoing cataract surgery were measured by high performance liquid chromatography (HPLC). METHODS: Totally 33 volunteer patients who scheduled for phacoemulsification surgery received one drop of levofloxacin every 6h for 3d before and on the day of surgery, administration of drug was stopped 1h before surgery. Levofloxacin concentration in aqueous humor was measured by HPLC method with fluorescence detector. RESULTS: A simple, effective and sensitive HPLC method for determination of levofloxacin in human ocular aqueous was validated. Linearity was shown for levofloxacin concentration over a wide range of 1.95×10-3-1.50 μg/mL. The mean aqueous level of levofloxacin was 0.3399±0.03405 μg/mL. CONCLUSION: Results from the present study demonstrate that topical administration of levofloxacin 0.5% before cataract surgery with routine dose (one drop every 6h) unable to reach MIC90 for most common microorganism causing acute bacterial endophthalmitis.     

The relapse risk of AML patients undergoing autologous transplantation correlates with the stem cell mobilizing potential

Autologous stem cell transplantation (ASCT) is widely used to consolidate first remission in AML. We determined the significance of circulating CD34+ cells at the day of blood stem cell collection in 78 AML patients. Patients mobilizing more than 60,000 CD34+ cells/ml had shorter overall survival (OS; P=0.0274), shorter time to progression (TTP; P=0.0014), and a higher relapse rate (P=0.0177). High levels of CD34+ cells were an independent marker for shorter OS and TTP in a multivariate analysis. These data suggest that ASCT is associated with unfavorable outcome in AML patients with high levels of mobilized peripheral CD34+ cells.     

Base excision repair deficiency caused by polymerase beta haploinsufficiency: accelerated DNA damage and increased mutational response to carcinogens

The base excision repair pathway (BER) is believed to maintain genomic integrity by repairing DNA damage arising spontaneously or induced by oxidizing and alkylating agents. To establish the role of DNA polymerase beta (beta-pol) in BER and beta-pol-dependent BER in maintaining genomic stability, we have measured the impact of a gene-targeted disruption in the beta-pol gene on DNA repair capacity and on in vivo sensitivity to carcinogens. We have extensively phenotyped the DNA beta-pol heterozygous (beta-pol(+/-)) mouse as expressing approximately 50% less beta-pol mRNA and protein and as exhibiting an equivalent reduction in the specific activity of beta-pol. We measured BER activity by in vitro G:U mismatch and (8-OH)G:C repair and find that there is a significant reduction in the ability of extracts from beta-pol(+/-) mice to repair these types of DNA damage. In vivo, the beta-pol(+/-) mice sustain higher levels of DNA single-strand breaks as well as increased chromosomal aberrations as compared with beta-pol(+/+) littermates. Additionally, we show that reduction in beta-pol expression and BER activity results in increased mutagenicity of dimethyl sulfate as evidenced by a 2-fold increase in LacI mutation frequency. Importantly, the beta-pol(+/-) mice do not exhibit increased sensitivity to DNA damage induced by N-nitroso-N-ethylurea, ionizing radiation, or UV radiation, which induce damage processed by alternative repair pathways, demonstrating that this model is specifically a BER-deficient model.     

Difference in the seminal plasma protein expression in unexplained infertile men with successful and unsuccessful in vitro fertilisation outcome

Unexplained male infertility (UMI) is a condition in which routine semen analysis fails to detect subcellular sperm dysfunctions. In the present research, a comparative proteomics study of seminal plasma (SP) was conducted in men with unexplained infertility whose female partners had undergone in vitro fertilisation (IVF) treatment to find differences in the SP protein profile. Five UMI men with successful and eight with unsuccessful IVF outcome enrolled in this study. Two‐dimensional polyacrylamide gel electrophoresis (2D‐PAGE) technique was used for protein separation. The differentially expressed proteins were identified using mass spectrometry. Results indicated that at least two different protein spots, including clusterin and epididymal secretory protein E1, were over‐expressed (1.5‐ and 2‐fold change, respectively, p < 0.05) while prostate‐specific antigen was downregulated (0.3‐fold change, p < 0.05) in the successful group as compared with the unsuccessful group. Considering the role of all three identified proteins in the sperm quality, the results of the present study introduced these proteins as new candidate biomarkers for success of IVF in UMI couples.     

A human laboratory study of the effects of quetiapine on subjective intoxication and alcohol craving

Rationale  

The available treatments for alcoholism are only modestly effective, and patients vary widely in their treatment response. Quetiapine, an atypical antipsychotic medication with antagonist activity at D1 and D2, 5-HT_1A and 5-HT_2A, H₁, and α1 and α2 receptors was shown to promote abstinence, reduce drinking days, and reduce heavy drinking days in a 12-week double-blind placebo-controlled trial.