Evolutionary insights from profiling LINE-1 activity at allelic resolution in a single human genome.

Transposable elements have created the majority of the sequence in many genomes. In mammals, LINE-1 retrotransposons have been expanding for more than 100 million years as distinct, consecutive lineages; however, the drivers of this recurrent lineage emergence and disappearance are unknown. Most human genome assemblies provide a record of this ancient evolution, but fail to resolve ongoing LINE-1 retrotranspositions. Utilizing the human CHM1 long-read-based haploid assembly, we identified and cloned all full-length, intact LINE-1s, and found 29 LINE-1s with measurable in vitro retrotransposition activity. Among individuals, these LINE-1s varied in their presence, their allelic sequences, and their activity. We found that recently retrotransposed LINE-1s tend to be active in vitro and polymorphic in the population relative to more ancient LINE-1s. However, some rare allelic forms of old LINE-1s retain activity, suggesting older lineages can persist longer than expected. Finally, in LINE-1s with in vitro activity and in vivo fitness, we identified mutations that may have increased replication in ancient genomes and may prove promising candidates for mechanistic investigations of the drivers of LINE-1 evolution and which LINE-1 sequences contribute to human disease.

Novel Concept of Alpha Satellite Cascading Higher-Order Repeats (HORs) and Precise Identification of 15mer and 20mer Cascading HORs in Complete T2T-CHM13 Assembly of Human Chromosome 15.

Unraveling the intricate centromere structure of human chromosomes holds profound implications, illuminating fundamental genetic mechanisms and potentially advancing our comprehension of genetic disorders and therapeutic interventions. This study rigorously identified and structurally analyzed alpha satellite higher-order repeats (HORs) within the centromere of human chromosome 15 in the complete T2T-CHM13 assembly using the high-precision GRM2023 algorithm. The most extensive alpha satellite HOR array in chromosome 15 reveals a novel cascading HOR, housing 429 15mer HOR copies, containing 4-, 7- and 11-monomer subfragments. Within each row of cascading HORs, all alpha satellite monomers are of distinct types, as in regular Willard's HORs. However, different HOR copies within the same cascading 15mer HOR contain more than one monomer of the same type. Each canonical 15mer HOR copy comprises 15 monomers belonging to only 9 different monomer types. Notably, 65% of the 429 15mer cascading HOR copies exhibit canonical structures, while 35% display variant configurations. Identified as the second most extensive alpha satellite HOR, another novel cascading HOR within human chromosome 15 encompasses 164 20mer HOR copies, each featuring two subfragments. Moreover, a distinct pattern emerges as interspersed 25mer/26mer structures differing from regular Willard's HORs and giving rise to a 34-monomer subfragment. Only a minor 18mer HOR array of 12 HOR copies is of the regular Willard's type. These revelations highlight the complexity within the chromosome 15 centromeric region, accentuating deviations from anticipated highly regular patterns and hinting at profound information encoding and functional potential within the human centromere.

Evaluation of CRISPR/Cas9 exon-skipping vector for choroideremia using human induced pluripotent stem cell-derived RPE.

BACKGROUND: Exon-skipping is a powerful genetic tool, especially when delivering genes using an AAV-mediated full-length gene supplementation strategy is difficult owing to large length of genes. Here, we used engineered human induced pluripotent stem cells and artificial intelligence to evaluate clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9-based exon-skipping vectors targeting genes of the retinal pigment epithelium (RPE). The model system was choroideremia; this is an X-linked inherited retinal disease caused by mutation of the CHM gene. METHODS: We explored whether artificial intelligence detected differentiation of human OTX2, PAX6 and MITF (hOPM) cells, in which OTX2, PAX6 and MITF expression was induced by doxycycline treatment, into RPE. Plasmid encoding CHM exon-skipping modules targeting the splice donor sites of exons 6 were constructed. A clonal hOPM cell line with a frameshift mutation in exon 6 was generated and differentiated into RPE. CHM exon 6-skipping was induced, and the effects of skipping on phagocytic activity, cell death and prenylation of Rab small GTPase (RAB) were evaluated using flow cytometry, an in vitro prenylation assay and western blotting. RESULTS: Artificial intelligence-based evaluation of RPE differentiation was successful. Retinal pigment epithelium cells with a frameshift mutation in exon 6 showed increased cell death, reduced phagocytic activity and increased cytosolic unprenylated RABs only when oxidative stress was in play. The latter two phenotypes were partially rescued by exon 6-skipping of CHM. CONCLUSIONS: CHM exon 6-skipping contributed to RPE phagocytosis probably by increasing RAB38 prenylation under oxidative stress.

Effects of Chinese herbal medicines on growth performance, intestinal flora, immunity and serum metabolites of hybrid grouper (Epinephelus fuscoguttatus♀×Epinephelus lanceolatu♂).

The immunomodulatory roles of Chinese herbal Medicine (CHM) in aquatic animals have been well-recorded. However, how CHM impacts the intestinal microbiota and serum metabolism is not fully understood. In this study, the effects of different additive levels of CHM on the growth performance, immunity, intestinal flora and serum metabolism of hybrid grouper (♀Epinephelus fuscoguttatus × â™‚Epinephelus lanceolatu) were investigated. The addition of 0.5%, 1.0%, 1.5% and 2.0% Chinese herbal medicine compound to feed could significantly improve the weight gain rate (WGR), specific growth rate (SGR) and survival rate (SR) of grouper, reduced feed coefficient, while had no significant difference on morphometric parameter. The most significant improvement for the parameters above was observed in 1.5% group. Different addition levels of CHM could also significantly enhance the activities of ACP, AKP, SOD, CAT and LZM in serum. Accordingly, the supplementation of CHM significantly induced up-regulation of immune genes such as IL-8, IL-1ß, TNF-α, Nrf2, Lzm in the liver, spleen and head kidney of grouper, improved the resistance of grouper to V. harveyi as well. The intestinal flora analysis showed that at the phylum level, the main dominant species of intestinal microorganisms were Firmicutes, Proteobacteria, Bacteroidota, Actinobacteriota, Gemmatimonadota, Desulfobacterota, Fusobacteriota and Myxococcota. At the genus level, the high abundance was Lactobacillus, Streptococcus, Bacteroides, Escherichia, Romboutsia, Sphingomonas and Muribaculaceae. The abundance of probiotics (such as Lachnospiraceae, Lactobacillaceae, Streptococcaceae, etc) in CHM-supplement groups were higher (highest in 1.5% group) compared with control group. Moreover, a total of 11 common differential metabolic pathways were screened by LC-MS metabolism analysis of serum, they were Neuroactive ligand-receptor interaction, Purine metabolism, Linoleic acid, Glycerophospholipid metabolism, Taurine and hypotaurine metabolism, Arginine and proline metabolism, ABC transporters, Aminoacyl-tRNA biosynthesis, Arachidonic acid metabolism, Drug metabolism-cytochrome P450, alpha-Linolenic acid metabolism. Also, three common differential metabolites (PI(20:4(5Z,8Z,11Z,14Z)/18:1(11Z)), PC(20:3(8Z,11Z,14Z)/22:1(13Z)), PC(22:0/20:4(5Z,8Z,11Z,14Z)) associated with intestinal health, growth and disease resistance was found. These data will contributes to a comprehensive understand for the regulatory roles of CHM on fish, which is also beneficial for the disease control and sustainable development of aquaculture.

Glycyrrhizin as a promising kryptonite against SARS-CoV-2: Clinical, experimental, and theoretical evidences.

COVID-19 is the most devastating disease in recent times affecting most people globally. The higher rate of transmissibility and mutations of SARS-CoV-2 along with the lack of potential therapeutics has made it a global crisis. Potential molecules from natural sources could be a fruitful remedy to combat COVID-19. This systematic review highlights the detailed therapeutic implication of naturally occurring glycyrrhizin and its related derivatives against COVID-19. Glycyrrhizin has already been established for blocking different biomolecular targets related to the SARS-CoV-2 replication cycle. In this article, several experimental and theoretical evidences of glycyrrhizin and related derivatives have been discussed in detail to evaluate their potential as a promising therapeutic strategy against COVID-19. Moreover, the implication of glycyrrhizin in traditional Chinese medicines for alleviating the symptoms of COVID-19 has been reviewed. The potential role of glycyrrhizin and related compounds in affecting various stages of the SARS-CoV-2 life cycle has also been discussed in detail. Derivatization of glycyrrhizin for designing potential lead compounds along with combination therapy with other anti-SARS-CoV-2 agents followed by extensive evaluation may assist in the formulation of novel anti-coronaviral therapy for better treatment to combat COVID-19.

Overexpression of Pericentromeric HSAT2 DNA Increases Expression of EMT Markers in Human Epithelial Cancer Cell Lines.

Pericentromeric tandemly repeated DNA of human satellites 1, 2, and 3 (HS1, HS2, and HS3) is actively transcribed in some cells. However, the functionality of the transcription remains obscure. Studies in this area have been hampered by the absence of a gapless genome assembly. The aim of our study was to map a transcript that we have previously described as HS2/HS3 on chromosomes using a newly published gapless genome assembly T2T-CHM13, and create a plasmid overexpressing the transcript to assess the influence of HS2/HS3 transcription on cancer cells. We report here that the sequence of the transcript is tandemly repeated on nine chromosomes (1, 2, 7, 9, 10, 16, 17, 22, and Y). A detailed analysis of its genomic localization and annotation in the T2T-CHM13 assembly revealed that the sequence belonged to HSAT2 (HS2) but not to the HS3 family of tandemly repeated DNA. The transcript was found on both strands of HSAT2 arrays. The overexpression of the HSAT2 transcript increased the transcription of the genes encoding the proteins involved in the epithelial-to-mesenchymal transition, EMT (SNAI1, ZEB1, and SNAI2), and the genes that mark cancer-associated fibroblasts (VIM, COL1A1, COL11A1, and ACTA2) in cancer cell lines A549 and HeLa. Co-transfection of the overexpression plasmid and antisense nucleotides eliminated the transcription of EMT genes observed after HSAT2 overexpression. Antisense oligonucleotides also decreased transcription of the EMT genes induced by tumor growth factor beta 1 (TGFß1). Thus, our study suggests HSAT2 lncRNA transcribed from the pericentromeric tandemly repeated DNA is involved in EMT regulation in cancer cells.

Choroideremia: The Endpoint Endgame.

Choroideremia is an X-linked retinal degeneration resulting from the progressive, centripetal loss of photoreceptors and choriocapillaris, secondary to the degeneration of the retinal pigment epithelium. Affected individuals present in late childhood or early teenage years with nyctalopia and progressive peripheral visual loss. Typically, by the fourth decade, the macula and fovea also degenerate, resulting in advanced sight loss. Currently, there are no approved treatments for this condition. Gene therapy offers the most promising therapeutic modality for halting or regressing functional loss. The aims of the current review are to highlight the lessons learnt from clinical trials in choroideremia, review endpoints, and propose a future strategy for clinical trials.

Immunohistochemical expression of BCL-2 in hydatidiform moles: a tissue microarray study.

Background: Hydatidiform moles (HM) are members of gestational trophoblastic diseases (GTD) and, in some cases, might progress to gestational trophoblastic neoplasia (GTN). HMs are either partial (PHM) or complete (CHM). Some HMs are challenging in arriving at a precise histopathological diagnosis. This study aims to investigate the expression of BCL-2 by immunohistochemistry (IHC) in HMs as well as in normal trophoblastic tissues "products of conception (POC) and placentas" using Tissue MicroArray (TMA) technique. Methods: TMAs were constructed using the archival material of 237 HMs (95 PHM and 142 CHM) and 202 control normal trophoblastic tissues; POC and unremarkable placentas. Sections were immunohistochemically stained using antibodies against BCL-2. The staining was assessed semi-quantatively (intensity and percentage of the positive cells) in different cellular components (trophoblasts and stromal cells). Results: BCL-2 showed cytoplasmic expression in more than 95% of trophoblasts of PHM, CHM and controls. The staining showed a significant reduction of the intensity from controls (73.7%), PHMs (76.3%) to CHM (26.9%). There was a statistically significant difference between PHM and CHM in the intensity (p-value 0.0005) and the overall scores (p-value 0.0005), but not the percentage score (p-value > 0.05). No significant difference was observed in the positivity of the villous stromal cells between the different groups. All cellular components were visible using the TMA model of two spots/case (3 mm diameter, each) in more than 90% of cases. Conclusions: Decreased BCL-2 expression in CHM compared to PHM and normal trophoblasts indicates increased apoptosis and uncontrolled trophoblastic proliferation. Construction of TMA in duplicates using cores of 3 mm diameter can overcome tissue heterogeneity of complex lesions.

Hydatidiform mole.

Hydatidiform mole is the most common form of gestational trophoblastic disease. It is an abnormally formed placental tissue with characteristic changes in karyotype, arising in fertilization disorders. The presence of abundant paternal genetic information plays a key role in the pathogenesis of complete and partial hydatidiform moles. These lesions are characterized by a relatively wide spectrum of morphological changes that may not be fully expressed, especially in the early stages of pregnancy. In addition, some changes can be observed in non-molar gravidities, which, unlike hydatidiform moles, lack any risk of malignant transformation. Although conventional histological examination still plays a key role in the diagnosis, it should be supplemented by other methods that reliably differentiate individual lesions. Accurate diagnosis of molar gravidities is important not only for determining the correct therapeutic approach, but the obtained data may also contribute to further research of these pathological entities.

Use of High-Resolution Multispectral UAVs to Calculate Projected Ground Area in Corylus avellana L. Tree Orchard.

In the last decade, research on Corylus avellana has focused on improving field techniques and hazelnut quality; however, climatic change and sustainability goals call for new agronomic management strategies. Precision management technologies could help improve resource use efficiency and increase grower income, but research on remote sensing systems and especially on drone devices is still limited. Therefore, the hazelnut is still linked to production techniques far from the so-called Agriculture 4.0. Unmanned aerial vehicles platforms are becoming increasingly available to satisfy the demand for rapid real-time monitoring for orchard management at spatial, spectral, and temporal resolutions, addressing the analysis of geometric traits such as canopy volume and area and vegetation indices. The objective of this study is to define a rapid procedure to calculate geometric parameters of the canopy, such as canopy area and height, by methods using NDVI and CHM values derived from UAV images. This procedure was tested on the young Corylus avellana tree to manage a hazelnut orchard in the early years of cultivation. The study area is a hazelnut orchard (6.68 ha), located in Bernalda, Italy. The survey was conducted in a six-year-old irrigated hazelnut orchard of Tonda di Giffoni and Nocchione varieties using multispectral UAV. We determined the Projected Ground Area and, on the Corylus avellana canopy trough, the vigor index NDVI (Normalized Difference Vegetation Index) and the CHM (Canopy Height Model), which were used to define the canopy and to calculate the tree crown area. The projection of the canopy area to the ground measured with NDVI values > 0.30 and NDVI values > 0.35 and compared with CHM measurements showed a statistically significant linear regression, R2 = 0.69 and R2 = 0.70, respectively. The ultra-high-resolution imagery collected with the UAV system helped identify and define each tree crown individually from the background (bare soil and grass cover). Future developments are the construction of reliable relationships between the vigor index NDVI and the Leaf Area Index (LAI), as well as the evaluation of their spatial-temporal evolution.

Genomic imprinting in human placentation.

Background: Genomic imprinting (GI) is a mammalian-specific epigenetic phenomenon that has been implicated in the evolution of the placenta in mammals. Methods: Embryo transfer procedures and trophoblast stem (TS) cells were used to re-examine mouse placenta-specific GI genes. For the analysis of human GI genes, cytotrophoblast cells isolated from human placental tissues were used. Using human TS cells, the biological roles of human GI genes were examined. Main findings: (1) Many previously identified mouse GI genes were likely to be falsely identified due to contaminating maternal cells. (2) Human placenta-specific GI genes were comprehensively determined, highlighting incomplete erasure of germline DNA methylation in the human placenta. (3) Human TS cells retained normal GI patterns. (4) Complete hydatidiform mole-derived TS cells were characterized by aberrant GI and enhanced trophoblastic proliferation. The maternally expressed imprinted gene p57KIP2 may be responsible for the enhanced proliferation. (5) The primate-specific microRNA cluster on chromosome 19, which is a placenta-specific GI gene, is essential for self-renewal and differentiation of human TS cells. Conclusion: Genomic imprinting plays diverse and important roles in human placentation. Experimental analyses using TS cells suggest that the GI maintenance is necessary for normal placental development in humans.

CHM mutation spectrum and disease: An update at the time of human therapeutic trials.

Choroideremia is an X-linked inherited retinal disorder (IRD) characterized by the degeneration of retinal pigment epithelium, photoreceptors, choriocapillaris and choroid affecting males with variable phenotypes in female carriers. Unlike other IRD, characterized by a large clinical and genetic heterogeneity, choroideremia shows a specific phenotype with causative mutations in only one gene, CHM. Ongoing gene replacement trials raise further interests in this disorder. We describe here the clinical and genetic data from a French cohort of 45 families, 25 of which carry novel variants, in the context of 822 previously reported choroideremia families. Most of the variants represent loss-of-function mutations with eleven families having large (i.e. ≥6 kb) genomic deletions, 18 small insertions, deletions or insertion deletions, six showing nonsense variants, eight splice site variants and two missense variants likely to affect splicing. Similarly, 822 previously published families carry mostly loss-of-function variants. Recurrent variants are observed worldwide, some of which linked to a common ancestor, others arisen independently in specific CHM regions prone to mutations. Since all exons of CHM may harbor variants, Sanger sequencing combined with quantitative polymerase chain reaction or multiplex ligation-dependent probe amplification experiments are efficient to achieve the molecular diagnosis in patients with typical choroideremia features.

Scheimpflug analysis of corneal power changes after hyperopic small incision lenticule extraction.

PURPOSE: To assess the ability of the Pentacam in predicting the corneal power after hyperopic small-incision lenticule extraction (SMILE). METHODS: Twenty-five eyes of 22 patients underwent hyperopic SMILE were prospectively followed. All patients finished at least 6 months visit. Cornea power was obtained by Pentacam HR, in the format of mean keratometry (Km), equivalent keratometry (EKR) and total cornea refractive power (TCRP). Calculation of TCRP were centered on either the corneal apex or the pupil center within a ring or zone, giving a total of four different subtypes naming AR、AZ、PR、PZ. Clinical history method (CHM) was regarded as a gold standard and was compared with other cornea power parameters. RESULTS: Center difference had no impact on the TCRP values (PR vs AR and PZ vs AZ, P > 0.05). Compared with CHM, no difference was found in Km, EKR 4.0 mm, EKR 4.5 mm, PR 3.0 mm, PR 4.0 mm, AR 3.0 mm and AR 4.0 mm. PR 4.0 mm showed the least difference with CHM (- 0.14 ± 1.03D, P > 0.05). The 95% limit of agreement (LOA) of the TCRPs and CHM was not close. The top two were PR 3.0 mm and PR 4.0 mm, LOA of which were - 2.20 to 1.84 D and - 2.18 to 1.68 D respectively. Central cornea thickness was correlated with error (TCRP - CHM) of PR 4.0 mm (r = 0.58, P = 0.003). CONCLUSIONS: The Pentacam topographer is an alternative method of measuring corneal power in eyes after hyperopic SMILE. The optimal options seem to be the TCRP (PR 4.0 mm). The agreement needs more verifications.

Scots pine stands biomass assessment using 3D data from unmanned aerial vehicle imagery in the Chernobyl Exclusion Zone.

Thirty-five years after the accident, large forest areas in the Chernobyl Exclusion Zone still contain huge amounts of radionuclides released from the Chernobyl Nuclear Power Plant Unit 4 in April 1986. An assessment of the radiological and radioecological consequences of persistent radioactive contamination and development of remediation strategies for Chernobyl forests imply acquiring comprehensive data on their contamination levels and dynamics of biomass inventories. The most accurate forest inventory data can be obtained in ground timber cruises. However, such cruises in radioactive contaminated forest ecosystems in the Chernobyl Exclusion Zone result in radiation exposures of the personnel involved, which means the need for development of the remote sensing methods. The purpose of this study is to analyze the applicability and limitations of the photogrammetric method for the remote large-scale monitoring of aboveground biomass inventories. Based on field measurements, we estimated the biomass inventories in 31 Scots pine stands including both artificial plantations and natural populations. The stands differed significantly in age (from a few years in natural populations to 115 years in the oldest plantation), productivity (from 0.4 to 19.8 kg m-2), mean height (from 4.1 to 36 m), and other parameters. Photogrammetric data were obtained from the same stands using unmanned aerial vehicle (UAV). These data were then processed using two approaches to derive the canopy height model (CHM) parameters which were tested for correlation with the aboveground biomass inventories. In the first approach, we found that the inventories correlated well with the mean value of CHM of the site (R2 = 0.79). In the second approach, the total aboveground biomass was approximated by a function of the average height of trees detected at the site and the total crown projection area (R2 = 0.78). Among other local parameters, the total crown projection area was identified as the major factor impacting the accuracy of the aboveground biomass inventory estimates from the UAV survey data in both approaches. In the dense stands with the high total crown projections areas (more than 0.90), the average relative deviations of the UAV-based aboveground biomass estimates from the results of the field measurements were close to 0, which means the adequate accuracy of the UAV surveys data for radioecological monitoring purposes. The relative deviations of the UAV-based estimates in both approaches increased in the stands consisting of separated groups of trees, which indicates potential limitation of the approaches and need for their further development.

Using canopy height model derived from UAV imagery as an auxiliary for spectral data to estimate the canopy cover of mixed broadleaf forests.

Canopy cover is an important structural trait that is frequently used in forest inventories to assess sustainability as well as many other important aspects of forest stands. Remote sensing data is more effective and suitable for canopy cover estimating than traditional field measurements such as sample plots, especially at broad scales. Measurement and mapping this attribute in fine-scale is a difficult task. Aerial imagery using unmanned aerial vehicle (UAV) has been recognized as an excellent tool to estimate canopy attributes. In this study, we compared the potential of using digital hemispherical photography (DHP), digital cover photography (DCP), UAV RGB data, and canopy height model (CHM) for estimation of canopy cover of mix broad-leaved forest on seven different stands. The canopy cover was measured from two digital canopy photographic methods, including DHP (as the reference method) and DCP. The stand orthophotos were segmented using a multi-resolution image segmentation method. Afterward, the classification in two classes of the canopy cover and the non-canopy cover was conducted using minimum distance classification to estimate canopy cover. The CHM layer was generated based on the SfM algorithm and utilized in the canopy cover estimation in each stand as auxiliary data. The results showed a slight improvement when we used the CHM as auxiliary data. Overall, the results showed that the efficiency of the ground digital canopy photographic methods (zenith view) in multi-storied and dense forests is the lowest. In return, our method for digital aerial canopy photography (object-based canopy segmentation and classification) is simple, quick, efficient, and cost-effective.

Balneotherapy with Chinese herbal medicine prolongs the remission period in patients with psoriasis vulgaris.

This study aimed to evaluate whether the supplementary balneotherapy with Chinese herbal medicine (CHM) could facilitate the treatment of psoriasis vulgaris and thus be beneficial for long-term remission from the symptoms. Two hundred psoriasis vulgaris patients with moderate-to-severe plaque psoriasis from January 2013 to June 2014 were evenly divided into two groups: the consolidated therapy group (CTG) and unconsolidated therapy group (UTG); the remission period of the two groups was compared. There was no significant difference in Psoriasis Area Severity Index (PASI) score between the two groups at the beginning and the end of the treatment. However, the average remission time in CTG was 10.99 months, which was significantly longer than that of 7.94 months in UTG (P = .001). After a correction of age, course of disease, skin type as well as PASI baseline value using a COX model, we found that the risk of recurrence of psoriasis vulgaris in UTG was higher than that in the CTG (P < .001). No adverse reactions were discovered when combing the two treatments together. The combined treatment of CHM balneotherapy and narrowband ultraviolet B could significantly prolong the remission time in patients with psoriasis vulgaris.

Next-generation sequencing-based clinical diagnosis of choroideremia and comprehensive mutational and clinical analyses.

BACKGROUND: To report the clinical and genetic findings from seven Chinese patients with choroideremia. METHODS: Five hundred seventy-eight patients with a clinically suspected diagnosis of retinitis pigmentosa (RP) underwent comprehensive ophthalmic examinations. Next-generation sequencing (NGS) was performed on samples from all patients. Detailed clinical characteristics of the patients with choroideremia identified in this study were assessed using multimodal imaging. RESULTS: Seven patients with choroideremia were identified, and six novel variants in CHM (c.1960 T > C p.Ter654Gln, c.1257del p.Ile420*fs1, c.1103_1121delATGGCAACACTCCATTTTT p.Tyr368Cysfs35, c.1414-2A > T, and c.1213C > T p.Gln405Ter, c.117-1G > A) were revealed. All variants were deleterious mutations: two were frameshifts, two were nonsense mutations, two were splicing mutations, and one was a readthrough mutation. The clinical phenotypes of these patients were markedly heterogeneous, and they shared many common clinical features with RP, including night blindness, constriction of the visual field and gradually reduced visual acuity. However, patients with choroideremia showed pigment hypertrophy and clumping, and chorioretinal atrophy, and a majority of patients with choroideremia presented with retinal tubulations in the outer layer of the retina. CONCLUSIONS: We provide a detailed description of the genotypes and phenotypes of seven patients with choroideremia who were accurately diagnosed using NGS. These findings provide a better understanding of the genetics and phenotypes of choroideremia.

Potential mechanisms of Chinese Herbal Medicine that implicated in the treatment of COVID-19 related renal injury.

Clinical studies have shown that renal injury in Corona Virus Disease 2019 (COVID-19) patients has been a real concern, which is associated with high mortality and an inflammation/apoptosis-related causality. Effective target therapy for renal injury has yet been developed. Besides, potential anti-COVID-19 medicines have also been reported to cause adverse side effects to kidney. Chinese Herbal Medicine (CHM), however, has rich experience in treating renal injury and has successfully applied in China in the battle of COVID-19. Nevertheless, the molecular mechanisms of CHM treatment are still unclear. In this study, we searched prescriptions in the treatment of renal injury extensively and the potential mechanisms to treat COVID-19 related renal injury were investigated. The association rules analysis showed that the core herbs includes Huang Qi, Fu Ling, Bai Zhu, Di Huang, Shan Yao. TCM herbs regulate core pathways, such as AGE-RAGE, PI3K-AKT, TNF and apoptosis pathway, etc. The ingredients (quercetin, formononetin, kaempferol, etc.,) from core herbs could modulate targets (PTGS2 (COX2), PTGS1 (COX1), IL6, CASP3, NOS2, and TNF, etc.), and thereby prevent the pharmacological and non-pharmacological renal injury comparable to that from COVID-19 infection. This study provides therapeutic potentials of CHM to combat COVID-19 related renal injury to reduce complications and mortality.

Pathogenicity of novel atypical variants leading to choroideremia as determined by functional analyses.

Choroideremia is a monogenic X-linked recessive chorioretinal disease linked to pathogenic variants in the CHM gene. These variants are commonly base-pair changes, frameshifts, or large deletions. However, a few rare or unusual events comprising large duplications, a retrotransposon insertion, a pseudo-exon activation, and two c-98 promoter substitutions have also been described. Following an exhaustive molecular diagnosis, we identified and characterized three novel atypical disease-causing variants in three unrelated male patients. One is a first-ever reported Alu insertion within CHM and the other two are nucleotide substitutions, c.-90C>G and c.-108A>G, affecting highly conserved promoter positions. RNA analysis combined with western blot and functional assays of patient cells established the pathogenicity of the Alu insertion and the c.-90C>G alteration. Furthermore, luciferase reporter assays suggested a CHM transcription defect associated with the c.-90C>G and c.-108A>G variants. These findings broaden our knowledge of the mutational spectrum and the transcriptional regulation of the CHM gene.

Optical coherence tomography (OCT) features of cystoid spaces in choroideremia (CHM).

PURPOSE: To investigate the prevalence and features of cystoid spaces (CS) in patients with confirmed genetic diagnosis of choroideremia (CHM) using swept source optical coherence tomography (OCT). METHODS: We retrospectively reviewed CHM patients examined at the Regional Reference Center for Hereditary Retinal Degenerations at the Eye Clinic in Florence. We took into consideration genetically confirmed CHM patients with ophthalmological and swept source optical coherence tomography (OCT) examinations. The presence/absence and location of cystoid spaces in the retina of each eye were reported. RESULTS: A total of 42 eyes of 21 CHM patients were included in our series. The average age of the patients was 36.5 ± 20.1 (range, 13-73 years). The average best-corrected visual acuity (BCVA) for all patients was 0.63 ± 1.00 logMar (range, 0-2,80). CS were present in 15 eyes of eight patients (8/21, 38%). In all cases, CS were located in inner nuclear layer (INL); in five eyes of three patients, CS were detected also in ganglion cell layer (GCL). CS appeared as microcistoyd abnormalities and were detected in retinal areas characterized by retinal pigment epithelium (RPE) and outer retinal layers atrophy at the transition zone. CONCLUSIONS: Cystoid spaces in choroideremia showed peculiar features; they are clusters of small-size extrafoveal degenerative cysts mainly located in inner nuclear layer at the transition zone where outer retinal layers and RPE are severely damaged.