Instrument-based Tests for Measuring Anterior Chamber Cells in Uveitis: A Systematic Review

ABSTRACT

Purpose

New instrument-based techniques for anterior chamber (AC) cell counting can offer automation and objectivity above clinician assessment. This review aims to identify such instruments and its correlation with clinician estimates.     

Treatment of diabetic autonomic neuropathy with an aldose reductase inhibitor

To evaluate the effects of the aldose reductase inhibitor Ponalrestat (Statil) on diabetic autonomic neuropathy, a double-blind placebo controlled trial was carried out on a group of 34 diabetic patients with documented cardiac autonomic neuropathy. After a 4-week, placebo run-in period, patients were randomised for treatment with 600 mg Statil or placebo for another 24 weeks. Moreover, the reliability of the autonomic nerve function tests was investigated by comparing the results at onset and at week 4. Fifteen patients treated with Statil and 12 with placebo completed the study. Neither symptom scores nor cardiovascular reflexes, pupil reflexes and skin vasomotor reflexes improved after Statil therapy, which led us to conclude that Statil is not effective in the treatment of diabetic autonomic neuropathy. Reliability coefficients for cardiovascular reflexes and pupil reflex showed high values, ranging from 60% to 80%. Therefore these methods are recommended in future therapy trials.     

A double cryptic chromosome imbalance is an important factor to explain phenotypic variability in Wolf-Hirschhorn syndrome

A total of five Wolf-Hirschhorn syndrome (WHS) patient with a 4p16.3 de novo microdeletion was referred because of genotype-phenotype inconsistencies, first explained as phenotypic variability of the WHS. The actual deletion size was found to be about 12 Mb in three patients, 5 Mb in another one and 20 Mb in the last one, leading us to hypothesize the presence of an extrachromosome segment on the deleted 4p. A der(4)(4qter --> p16.1::8p23 --> pter) chromosome, resulting from an unbalanced de novo translocation was, in fact, detected in four patients and a der(4)(4qter --> q32::4p15.3 --> qter) in the last. Unbalanced t(4;8) translocations were maternal in origin, the rec(4p;4q) was paternal. With the purpose of verifying frequency and specificity of this phenomenon, we investigated yet another group of 20 WHS patients with de novo large deletions (n = 13) or microdeletions (n = 7) and with apparently straightforward genotype-phenotype correlations. The rearrangement was paternal in origin, and occurred as a single anomaly in 19 out of 20 patients. In the remaining patient, the deleted chromosome 4 was maternally derived and consisted of a der(4)(4qter --> 4p16.3::8p23 --> 8pter). In conclusions, we observed that 20% (5/25) of de novo WHS-associated rearrangements were maternal in origin and 80% (20/25) were paternal. All the maternally derived rearrangements were de novo unbalanced t(4;8) translocations and showed specific clinical phenotypes. Paternally derived rearrangements were usually isolated deletions. It can be inferred that a double, cryptic chromosome imbalance is an important factor for phenotypic variability in WHS. It acts either by masking the actual deletion size or by doubling a quantitative change of the genome.     

Complex mosaic structural variations in human fetal brains

Somatic mosaicism, manifesting as single nucleotide variants (SNVs), mobile element insertions, and structural changes in the DNA, is a common phenomenon in human brain cells, with potential functional consequences. Using a clonal approach, we previously detected 200–400 mosaic SNVs per cell in three human fetal brains (15–21 wk postconception). However, structural variation in the human fetal brain has not yet been investigated. Here, we discover and validate four mosaic structural variants (SVs) in the same brains and resolve their precise breakpoints. The SVs were of kilobase scale and complex, consisting of deletion(s) and rearranged genomic fragments, which sometimes originated from different chromosomes. Sequences at the breakpoints of these rearrangements had microhomologies, suggesting their origin from replication errors. One SV was found in two clones, and we timed its origin to ∼14 wk postconception. No large scale mosaic copy number variants (CNVs) were detectable in normal fetal human brains, suggesting that previously reported megabase-scale CNVs in neurons arise at later stages of development. By reanalysis of public single nuclei data from adult brain neurons, we detected an extrachromosomal circular DNA event. Our study reveals the existence of mosaic SVs in the developing human brain, likely arising from cell proliferation during mid-neurogenesis. Although relatively rare compared to SNVs and present in ∼10% of neurons, SVs in developing human brain affect a comparable number of bases in the genome (∼6200 vs. ∼4000 bp), implying that they may have similar functional consequences.

Somatic mosaicism, the presence of more than one genotype in the somatic cells of an individual, is a prominent phenomenon in the human central nervous system. Forms of mosaicism include aneuploidies and smaller copy number variants (CNVs), structural variants (SVs), mobile element insertions, indels, and single nucleotide variants (SNVs). The developing human brain exhibits high levels of aneuploidy compared to other tissues, generating genetic diversity in neurons (Pack et al. 2005; Yurov et al. 2007; Bushman and Chun 2013). Such aneuploidy was suggested to be a natural feature of neurons, rather than a distinctive feature of neurodegeneration. However, the frequency of aneuploidy in neurons has been debated, with a separate study suggesting that aneuploidies occur in only about 2.2% of mature adult neurons (Knouse et al. 2014). They hence infer that such aneuploidy could have adverse effects at the cellular and organismal levels. Additionally, analysis of single cells from normal and pathological human brains identified large, private, and likely clonal somatic CNVs in both normal and diseased brains (Gole et al. 2013; McConnell et al. 2013; Cai et al. 2014; Knouse et al. 2016; Chronister et al. 2019; Perez-Rodriguez et al. 2019), with 3%–25% of human cerebral cortical nuclei carrying megabase-scale CNVs (Chronister et al. 2019) and deletions being twice as common as duplications (McConnell et al. 2013). Given that CNVs often arise from nonhomologous recombination and replication errors, their likely time of origin is during brain development. However, when CNVs first arise in human brain development has not yet been investigated. The present work is the first to examine this question using clonal populations of neuronal progenitor cells (NPCs) obtained from fetal human brains.Detection of CNVs in single neurons is challenging, given the need to amplify DNA. Such amplification may introduce artifacts that could, in turn, be misinterpreted as CNVs. In order to address this technical limitation, Hazen et al. reprogrammed adult postmitotic neurons using somatic cell nuclear transfer (SCNT) of neuronal nuclei into enucleated oocytes (Hazen et al. 2016). These oocytes then made sufficient copies of the neuronal genome allowing for whole-genome sequencing (WGS), thus eliminating the need for amplification in vitro. Using this method, they identified a total of nine structural variants in six neurons from mice, three of which were complex rearrangements. However, it is not possible to extend such studies to humans, given the ethical issues involved, besides the technical challenges in obtaining and cloning adult neurons. To circumvent the need of single-cell DNA amplification or nuclear cloning, we examined clonal cell populations obtained from neural progenitor cells from the frontal region of the cerebral cortex (FR), parietal cortex (PA) and basal ganglia (BG) and describe here the discovery and analysis of mosaic SVs in these NPCs (Bae et al. 2018). These clones were sequenced at 30× coverage (much higher than most previous single-cell studies), allowing identification of SVs other than large deletions and duplications as well as precise breakpoint resolution.     

Antibiotic resistance in exopolysaccharide-forming Staphylococcus epidermidis clinical isolates from orthopaedic implant infections

The opportunistic pathogen Staphylococcus epidermidis is able to produce biofilm and to frequently cause implant infections. In recent years, it has also exhibited an increasing antimicrobial drug resistance. Here, the resistance to a panel of 16 different antibiotics in 342 clinical strains of S. epidermidis from orthopaedic implant infections has been investigated. The isolates were pheno- and genotyped for extracellular polysaccharide production, relevant to staphylococcal biofilm formation, in order to ascertain possible associations with antibiotic resistance. Approximately 10% of the isolates were found to be sensitive to all screened antibiotics. In all, 37-38% were resistant to beta-lactams such as oxacillin and imipenem, while the resistance to penicillin, ampicillin, cefazolin, cefamandole, was consistently observed in over 80% of the strains. Erythromycin- and clindamycin- resistant strains were approximately 41% and 16%, respectively. Of the isolates, 10% was resistant to chloramphenicol, 23% to sulfamethoxazole and 26% to ciprofloxacin. Resistance to vancomycin was never observed. Interestingly, exopolysaccharide-producing strains exhibited a significantly higher prevalence in the resistance to the four aminoglycosides (gentamicin, amikacin, netilmicin, tobramycin), to sulfamethoxazole and to ciprofloxacin with respect to non-producing isolates. Moreover, multiple resistance to antibiotics was more frequent among exopolysaccharide-forming strains.     

Light scattering from aqueous solutions of poly(acrylamide-co-maleic anhydride)

Light scattering measurements from aqueous solutions for two samples of poly(acrylamide-co-maleic anhydride) in the presence of salt are conducted. One of the samples exhibits a negative initial slope and a minimum in the plot of the conventional reciprocal scattered intensity function of sin² (θ/2). The explanation for this anomaly is a large optical anisotropy of the segment. A correction for this effect of segmental anisotropy is made. The behaviour of the other sample of this copolymer is typical for a polyelectrolyte. By analysing the disymmetry of the scattered light, the influence of the salt concentration on the polyion is evidenced by the determination of the macromolecular dimensions. Light scattering data allow the evaluation of the Mark-Kuhn-Houwink exponent a for various degrees of dissociation of the polyion; a correlation between the variation of a and the shape of the macromolecule is observed.     

Early and late paternal contribution to cell division of embryos in a time-lapse imaging incubation system

The objective of this study was to investigate the impact of male age, semen quality and days of ejaculatory abstinence on embryo morphokinetics. A total of 1,220 zygotes obtained from 139 couples in a private in vitro fertilisation centre were analysed. The timing of specific events from the point of insemination, such as timings to pronuclei appearance and fading, to two, three, four, five, six, seven and eight cells and to blastulation were recorded. Multivariate linear regression analysis was used to evaluate the influence of paternal factors on embryo morphokinetic events. Paternal age was positively correlated with delayed cell cleavage and blastulation, and negatively associated with implantation rate, and clinical pregnancy and live–birth chances. The ejaculatory abstinence was inversely correlated with the implantation rate. Inverse relationships were observed between semen parameters (sperm count, progressive sperm motility, total motile sperm count and morphology) and the timing of specific events during embryo development. Sperm morphology was also positively associated with implantation rate and pregnancy and live–birth chances. Increased paternal age and ejaculatory abstinence, and poor semen quality correlate with delayed cell cleavage and blastulation and negatively impact intracytoplasmic sperm injection outcomes.     

Steriods for multiple sclerosis and optic neuritis: a meta-analysis of randomized controlled clinical trials

We conducted a meta-analysis of randomized controlled clinical trials on steroid treatment for multiple sclerosis and optic neuritis. Of the 25 trials comparing steroids and controls without steroid treatment that we identified 12 were selected for this review. A meta-analysis was conducted to calculate the overall odds ratio across the studies for the numbers of patients without functional improvement and with new relapses. The trials included a total of 1714 patients: 998 with multiple sclerosis and 716 with optic neuritis. Any type of corticosteroids or adrenocorticotropic hormone (ACTH) treatment was considered, as was any dosage, route of administration, and length of treatment. Main outcome measures were: (a) number of multiple sclerosis patients who did not improve by at least one point on the EDSS or equivalent scale, or number of optic neuritis patients without complete recovery of visual acuity at 8 or 30 days and at longer follow-up; (b) number of multiple sclerosis patients with at least one new relapse, or number of optic neuritis patients in whom definite multiple sclerosis was diagnosed at longer follow-up. We found that corticosteroids or ACTH produced a significant improvement in disability or visual acuity at 30 days (odds ratio 0.49; 95 % CI 0.37–0.64). The improvement was not statistically significant at longer follow-up (0.85; 95 % CI 0.67–1.09). The treatment did not significantly reduce the number of patients with relapses (0.74; 95 % CI 0.54–1.01). Both low and high doses were effective for 30-day improvement, but only high-dose and short-term therapy were factors that identified subgroups with some reduction in the risk of new relapse. However, the power of the statistical analysis to detect a reliable difference in the subgroups was low. Steroid treatment is therefore effective in accelerating short-term recovery in patients with multiple sclerosis or optic neuritis. Whether steroids are also effective in reducing the risk of relapse, and the optimal dose and length of treatment must still be determined. Received: 5 August 1999, Received in revised form: 29 December 1999, Accepted: 22 January 2000     

Safety and Efficacy of Eucaloric Very Low-Carb Diet (EVLCD) in Type 1 Diabetes: A One-Year Real-Life Retrospective Experience

A eucaloric very low carbohydrate diet (EVLCD) is a diet with a daily caloric intake equal to the total daily energy expenditure (TDEE) with a carbohydrate content of <50 g/day. The literature on very low carbohydrate diets (VLCD) in type 1 diabetes (DM 1) is limited, although recently published scientific studies have highlighted their safety and efficacy in managing DM 1. In this retrospective analysis, we report the clinical data of 33 patients affected by DM 1 carrying out insulin therapy who switched voluntarily from their usual diet (high carb, low fat) to an EVLCD. Our aim is to evaluate the glycemic control, the amount of insulin needed in order to maintain glycemic control and safety of EVLCD. The switch improved glycemic control (mean glycated hemoglobin decreased from 8.3% to 6.8% (p < 0.01). The number of patients who reached a glycated hemoglobin value of <7% increased statistically from 12% to 57% (p < 0.01), and there was a statistically significant decrease (p < 0.01) in the units of daily insulin (from 36.7± 14.9 IU to 28.9 ±9.1 IU) A reduction from 54% to 24% in clinical level 2 hypoglycemia episodes was reported. No cases of severe hypoglycemia or ketoacidosis were observed. The results of the study support that EVLCD in DM 1 seems safe and effective when adopted under tight medical supervision.     

捷克中医药发展现状与分析

捷克作为欧洲内陆的议会共和制国家,实施强制保险模式以保障民众健康。导致人口死亡的主要疾病为缺血性心脏病、脑卒中和肺癌,而糖尿病、AD及COPD近年来发病率显著增加。捷克的传统医学包括巴赫花疗法、特异性免疫调节自我疗法、法国耳针治疗等。中医药在捷克具有一定的接受度,尤其是针灸已在神经性疾病中得到广泛应用。但中草药在捷克的使用受到限制,且中医药执业尚未合法化。建议深化多层次、多领域交流合作,并通过加强高水平中医药学位教育,加大中药与非药物疗法宣传和普及力度,推动中医药在捷克的发展。