Synthesis and Characterization of a Luminol Based Chemiluminescent Trimeric System.

A luminol based chemiluminescent trimeric system, namely 2,3-dihydro-5,8-di(thiophen-2-yl)phthalazine-1,4-dione (T2B-Lum), bearing thiophene rings as donor units and 2,3-dihydrophthalazine-1,4-dione as an acceptor unit was synthesized in two steps via donor-acceptor-donor approach using two different methods. It was found that T2B-Lum emits chemiluminescent light when exposed to H2O2 in a basic medium, and the presence of substituents and the type of aromatic ring bearing chemiluminescent active group have a direct effect on the compound's sensitivity. Among the members of a large family of metal ions, fluorescent and chemiluminescent T2B-Lum exhibited high sensitivity to Cu2+ and Fe3+ ions. Except for other metal cations (silver(I), cadmium(II), cobalt(II), iron(III), lithium(I), magnesium(II), manganese(II), nickel(II), zinc(II)), it has been observed that T2B-Lum is mostly sensitive to copper(II) ions with a detection limit value of 2.2 × 10- 3 M. On the other hand, T2B-Lum was also found to exhibit a high sensitivity to extremely dilute aqueous solutions (e.g., 1:50.000 dilution) of blood samples, making it a promising candidate for use in forensic applications.

Short-term outcomes of pregnant women with convalescent COVID-19 and factors associated with false-negative polymerase chain reaction test: A prospective cohort study.

AIM: To evaluate the clinical factors associated with false-negative RT-PCR results and to report the outcome of a cohort of pregnant women with COVID-19. METHODS: This cohort study was conducted in a tertiary referral pandemic hospital and included 56 pregnant women. A study including pregnant women with either a laboratory or clinical diagnosis for COVID-19 were included in the study. The primary outcome was clinical factors associated with false-negative RT-PCR results defined as a positive immunoglobulin M assessed by rapid testing in clinically diagnosed patients. Clinical outcomes of laboratory diagnosed patients were also reported. RESULTS: In total, 56 women with either RT-PCR or clinical COVID-19 diagnosis were included in the study. Forty-three women either had RT-PCR positivity or IgM positivity. The clinical outcome of these pregnancies was as follows: mean maternal age 27.7, immunoglobulin M positive patients 76.7%, RT-PCR positive patients 55.8%, maternal comorbidities 11.5%, complications in patients below 20 weeks 34.8%, complications in patients above 20 weeks 65.1%, elevated CRP 83.7%, lymphopenia 30.2%, time from hospital admission to final follow-up days 37 and stillbirth 8.3%. The proportion of women who tested positive for SARS-CoV-2 immunoglobulin M was 100% in the RT-PCR positive group and 56.5% in the clinical diagnosis group (P = .002). The symptom onset to RT-PCR testing interval longer than a week (risk ratio: 2.72, 95% CI: 1.14-5.40, P = .003) and presence of dyspnoea (risk ratio: 0.38, 95% CI: 0.14-0.89, P = .035) were associated with false-negative RT-PCR tests. The area under the curve of these parameters predicting false-negative RT-PCR was 0.73 (95% CI: 0.57-0.89). CONCLUSIONS: Symptomatic women with a negative RT-PCR should not be dismissed as potential COVID-19 patients, especially in the presence of prolonged symptom onset-test interval and in women without dyspnoea.

Does Hypochlorous Acid Cause Ototoxicity? An Experimental Study.

AIM: Hypochlorous acid (HOCl) is a weak acid that ionizes in water. It is an effective antiseptic exhibiting low toxicity on living tissues. We aimed to investigate the ototoxic effects of HOCl on an animal model by using electrophysiological and histological methods. MATERIALS AND METHODS: The study comprised 32 Sprague-Dawley rats, which were separated into four groups: control group (A), saline solution group (B), 70% isopropyl alcohol + 2% chlorhexidine group (C), and HOCl group (D). After recording the auditory brainstem response (ABR) for basal hearing thresholds (8, 16, 24, and 32 kHz), 0.03 ml of the aforementioned materials was injected intratympanically three times every 2 days in groups B, C, and D. ABR measurements were repeated on the 7th and 21st days. All animals were sacrificed, and temporal bones were prepared for examinations of cochlear histology and vascular endothelial growth factor immunohistochemistry. RESULTS: Basal hearing levels were normal across all frequencies and groups, with no statistical differentiation. On the 7th and 21st days after the ABR test, all other groups demonstrated a significant deterioration in hearing levels compared with group A. When the results from 7th and 21st days were compared within group D, a partial recovery was observed. In histopathology, groups C and D demonstrated moderate and severe cochlear degeneration, along with decreased immunoreactivity in the organ of Corti, stria vascularis, and spiral ligament. CONCLUSION: This is the first study to evaluate the safety of using HOCl in otology. Although HOCI is less ototoxic than the disinfectant used, it may have a toxic effect on cochlea.Level of Evidence: Animal Research.

An autosomal recessive limb girdle muscular dystrophy (LGMD2) with mild mental retardation is allelic to Walker-Warburg syndrome (WWS) caused by a mutation in the POMT1 gene.

Mutations of the protein O-mannosyltransferase (POMT1) gene affect glycosylation of alpha-dystroglycan, leading to Walker-Warburg syndrome, a lethal disorder in early life with severe congenital muscular dystrophy, and brain and eye malformations. Recently, we described a novel form of recessive limb girdle muscular dystrophy with mild mental retardation, associated with an abnormal alpha-dystroglycan pattern in the muscle, suggesting a glycosylation defect. Here, we present evidence that this distinct phenotype results from a common mutation (A200P) in the POMT1 gene. Our findings further expand the phenotype of glycosylation disorders linked to POMT1 mutations. Furthermore, the A200P mutation is part of a conserved core haplotype, indicating an ancestral founder mutation.

Identification of an ancestral haplotype of the 35delG mutation in the GJB2 (connexin 26) gene responsible for autosomal recessive non-syndromic hearing loss in families from the Eastern Black Sea Region in Turkey.

Mutations in the GJB2 gene have been shown to be the major cause of autosomal recessively inherited, prelingual, non-syndromic hearing loss. 35delG was found to be the most frequent mutation among Caucasians. In this study, we performed haplotype analysis of two large families with autosomal recessive non-syndromic hearing loss (totally 33 affected, 37 unaffected) from Trabzon (a city from the Eastern Black Sea region) by using polymorphic markers close to the 35delG mutation region, and identified a common haplotype, "2-6-4". The frequency of the mutant chromosomes having the 2-6-4 haplotype was compared between the Eastern Black Sea region and the other regions of Turkey and the difference was found to be significant (chi squared = 5.13/df = 1/p = 0.023). Also, when the frequency of mutant and wild type chromosomes having the 2-6-4 haplotype was compared in the Eastern Black Sea region, a statistically significant difference was observed in the mutant chromosomes (chi squared = 7.46/df = 1/p < or = 0.01). The results of this study demonstrate that the ancestral haplotype of the chromosomes bearing 35delG mutation in the Eastern Black Sea region is "2-6-4".

A novel form of recessive limb girdle muscular dystrophy with mental retardation and abnormal expression of alpha-dystroglycan.

The limb girdle muscular dystrophies are a heterogeneous group of conditions characterized by proximal muscle weakness and disease onset ranging from infancy to adulthood. We report here eight patients from seven unrelated families affected by a novel and relatively mild form of autosomal recessive limb girdle muscular dystrophy (LGMD2) with onset in the first decade of life and characterized by severe mental retardation but normal brain imaging. Immunocytochemical studies revealed a significant selective reduction of alpha-dystroglycan expression in the muscle biopsies. Linkage analysis excluded known loci for both limb girdle muscular dystrophy and congenital muscular dystrophies in the consanguineous families. We consider that this represents a novel form of muscular dystrophy with associated brain involvement. The biochemical studies suggest that it may belong to the growing number of muscular dystrophies with abnormal expression of alpha-dystroglycan.

Efficient transfection of mouse-derived C2C12 myoblasts using a matrigel basement membrane matrix.

Myogenic cell lines have been used widely in the study of myogenic differentiation, muscle regeneration and homeostasis, but, myoblasts and myotubes are difficult to transfect using conventional techniques. We have used liposome-based transfection method to introduce a green fluorescence protein (GFP)-expressing plasmid into Matrigel basement membrane matrix-coated C2C12 mouse myoblast cells. Myoblasts adhered and proliferated more rapidly on a Matrigel; thus, a dramatic increase in transfection efficiency can be obtained compared to Matrigel-untreated cells. Transfection efficiency was determined by counting fluorescent and total cells from six random fields for each condition. This protocol results in efficient (up to 60-70%) transfection of C2C12 myoblasts, high levels of GFP expression and low rate of cell death (10%). This technique is rapid, reliable, uses a lipid-based transfection reagent, and yields high transfection rates in a previously hard-to-transfect cell type.

Prenatal diagnosis of muscle-eye-brain disease.

OBJECTIVES: To present a family in which it was possible to perform prenatal diagnosis for the recessively inherited muscle-eye-brain disease (MEB) using linkage analysis. METHODS: Linkage analysis and direct sequencing of the POMGNT1 gene were carried out in a Turkish MEB family with one affected individual. Fetal DNA was obtained from an ongoing pregnancy by chorionic villus sampling (CVS). RESULTS: Both linkage analysis of the POMGNT1/1p32-p34 region and direct sequencing for the novel familial mutation (R605H) demonstrated that the fetus did not have MEB. CONCLUSION: We report the first case of prenatal diagnosis in MEB by molecular genetic analysis.

Calpain-3 mutations in Turkey.

Autosomal recessive limb-girdle muscular dystrophies (LGMD2s) are a clinically and genetically heterogeneous group of disorders, characterized by progressive involvement of the proximal limb girdle muscles; the group includes at least 10 different genetic entities. The calpainopathies (LGMD2A), a subgroup of LGMD2s, are estimated to be the most common forms of LGMD2 in all populations so far investigated. LGMD2A is usually characterized by symmetrical and selective atrophy of pelvic, scapular and trunk muscles and a moderate to gross elevation of serum CK. However, the course is highly variable. It is caused by mutations in the CAPN3 gene, which encodes for the calpain-3 protein. Until now, 161 pathogenic mutations have been found in the CAPN3 gene. In the present study, through screening of 93 unrelated LGMD2 families, we identified 29 families with LGMD2A, 21 (22.6%) of which were identified as having CAPN3 gene mutations. We detected six novel (p.K211N, p.D230G, p.Y322H, p.R698S, p.Q738X, c.2257delGinsAA) and nine previously reported mutations (c.550delA, c.19_23del, c.1746-20C>G, p.R49H, p.R490Q, p.Y336N, p.A702V, p.Y537X, p.R541Q) in the CAPN3 gene. There may be a wide variety of mutations, but clustering of specific mutations (c.550delA: 40%, p.R490Q: 10%) could be used in the diagnostic scheme in Turkey.