Modeling antisense oligonucleotide therapy in MECP2 duplication syndrome human iPSC-derived neurons reveals gene expression programs responsive to MeCP2 levels.

Genomic copy-number variations (CNVs) that can cause neurodevelopmental disorders often encompass many genes, which complicates our understanding of how individual genes within a CNV contribute to pathology. MECP2 duplication syndrome (MDS or MRXSL in OMIM; OMIM#300260) is one such CNV disorder caused by duplications spanning methyl CpG-binding protein 2 (MECP2) and other genes on Xq28. Using an antisense oligonucleotide (ASO) to normalize MECP2 dosage is sufficient to rescue abnormal neurological phenotypes in mouse models overexpressing MECP2 alone, implicating the importance of increased MECP2 dosage within CNVs of Xq28. However, because MDS CNVs span MECP2 and additional genes, we generated human neurons from multiple MDS patient-derived induced pluripotent cells (iPSCs) to evaluate the benefit of using an ASO against MECP2 in a MDS human neuronal context. Importantly, we identified a signature of genes that is partially and qualitatively modulated upon ASO treatment, pinpointed genes sensitive to MeCP2 function, and altered in a model of Rett syndrome, a neurological disorder caused by loss of MeCP2 function. Furthermore, the signature contained genes that are aberrantly altered in unaffected control human neurons upon MeCP2 depletion, revealing gene expression programs qualitatively sensitive to MeCP2 levels in human neurons. Lastly, ASO treatment led to a partial rescue of abnormal neuronal morphology in MDS neurons. All together, these data demonstrate that ASOs targeting MECP2 benefit human MDS neurons. Moreover, our study establishes a paradigm by which to evaluate the contribution of individual genes within a CNV to pathogenesis and to assess their potential as a therapeutic target.

Impact of Interleukin-6 on Oral Squamous Cell Carcinoma Among the South Indian Population.

Introduction Oral squamous cell carcinoma (OSCC) is associated with high rates of morbidity and mortality. Despite advances in research and treatment, the survival rate of OSCC patients has not changed considerably in recent years. Interleukin-6 (IL-6) is a proinflammatory cytokine that is involved in the development of various cancers including OSCC. The role of IL-6 is being studied in various cancers; however, its exact mechanism of action in OSCC among the South Indian population has not yet been studied. Thus, the current study aims to evaluate and assess the impact of IL-6 on OSCC among the South Indian population. Materials and methods Twenty tissues from OSCC patients and 20 normal tissues surrounding the same area from normal people were gathered from the Department of Oral and Maxillofacial Surgery, Saveetha Dental College and Hospital. The tissues were prepared for expression investigations and hematoxylin and eosin staining. The data was presented as mean ± standard deviation, with statistical significance at p<0.05. Results Our results indicate that, in comparison to normal tissues, OSCC samples had increased IL-6 expression levels (p<0.05). Conclusion We conclude that IL-6 has been identified as a key oncogene in the development of tumors and their spread in several types of cancers, including OSCC. Therefore, IL-6 can be used as a potential diagnostic or prognostic biomarker and the use of IL-6 inhibitors can be formulated as a potential treatment for OSCC.

Optimized self-attention based cycle-consistent generative adversarial network adopted melanoma classification from dermoscopic images.

Skin is the exposed part of the human body that constantly protected from UV rays, heat, light, dust, and other hazardous radiation. One of the most dangerous illnesses that affect people is skin cancer. A type of skin cancer called melanoma starts in the melanocytes, which regulate the colour in human skin. Reducing the fatality rate from skin cancer requires early detection and diagnosis of conditions like melanoma. In this article, a Self-attention based cycle-consistent generative adversarial network optimized with Archerfish Hunting Optimization Algorithm adopted Melanoma Classification (SACCGAN-AHOA-MC-DI) from dermoscopic images is proposed. Primarily, the input Skin dermoscopic images are gathered via the dataset of ISIC 2019. Then, the input Skin dermoscopic images is pre-processed using adjusted quick shift phase preserving dynamic range compression (AQSP-DRC) for removing noise and increase the quality of Skin dermoscopic images. These pre-processed images are fed to the piecewise fuzzy C-means clustering (PF-CMC) for ROI region segmentation. The segmented ROI region is supplied to the Hexadecimal Local Adaptive Binary Pattern (HLABP) to extract the Radiomic features, like Grayscale statistic features (standard deviation, mean, kurtosis, and skewness) together with Haralick Texture features (contrast, energy, entropy, homogeneity, and inverse different moments). The extracted features are fed to self-attention based cycle-consistent generative adversarial network (SACCGAN) which classifies the skin cancers as Melanocytic nevus, Basal cell carcinoma, Actinic Keratosis, Benign keratosis, Dermatofibroma, Vascular lesion, Squamous cell carcinoma and melanoma. In general, SACCGAN not adapt any optimization modes to define the ideal parameters to assure accurate classification of skin cancer. Hence, Archerfish Hunting Optimization Algorithm (AHOA) is considered to maximize the SACCGAN classifier, which categorizes the skin cancer accurately. The proposed method attains 23.01%, 14.96%, and 45.31% higher accuracy and 32.16%, 11.32%, and 24.56% lesser computational time evaluated to the existing methods, like melanoma prediction method for unbalanced data utilizing optimized Squeeze Net through bald eagle search optimization (CNN-BES-MC-DI), hyper-parameter optimized CNN depending on Grey wolf optimization algorithm (CNN-GWOA-MC-DI), DEANN incited skin cancer finding depending on fuzzy c-means clustering (DEANN-MC-DI). RESEARCH HIGHLIGHTS: This manuscript, self-attention based cycle-consistent. SACCGAN-AHOA-MC-DI method is implemented in Python. (SACCGAN-AHOA-MC-DI) from dermoscopic images is proposed. Adjusted quick shift phase preserving dynamic range compression (AQSP-DRC). Removing noise and increase the quality of Skin dermoscopic images.

A novel pathogenic mutation of MeCP2 impairs chromatin association independent of protein levels.

Loss-of-function mutations in MECP2 cause Rett syndrome (RTT), a severe neurological disorder that mainly affects girls. Mutations in MECP2 do occur in males occasionally and typically cause severe encephalopathy and premature lethality. Recently, we identified a missense mutation (c.353G>A, p.Gly118Glu [G118E]), which has never been seen before in MECP2, in a young boy who suffered from progressive motor dysfunction and developmental delay. To determine whether this variant caused the clinical symptoms and study its functional consequences, we established two disease models, including human neurons from patient-derived iPSCs and a knock-in mouse line. G118E mutation partially reduces MeCP2 abundance and its DNA binding, and G118E mice manifest RTT-like symptoms seen in the patient, affirming the pathogenicity of this mutation. Using live-cell and single-molecule imaging, we found that G118E mutation alters MeCP2's chromatin interaction properties in live neurons independently of its effect on protein levels. Here we report the generation and characterization of RTT models of a male hypomorphic variant and reveal new insight into the mechanism by which this pathological mutation affects MeCP2's chromatin dynamics. Our ability to quantify protein dynamics in disease models lays the foundation for harnessing high-resolution single-molecule imaging as the next frontier for developing innovative therapies for RTT and other diseases.

Antidiabetic activity of Kabasura Kudineer Chooranam.

One of the traditional plants used in Siddha medicine is Kabasura Kudineer Chooranam. It is said to possess antiaging, life-strengthening, and disease-preventing activities that have an enormous influence on health care. It has significant therapeutic potential and ethnobotanical significance. The aim of this study is to investigate the antidiabetic activity of Kabasura Kudineer Chooranam. The antidiabetic potential of Kabasura Kudineer Chooranam was determined in vitro using established methods such as alpha-amylase and alpha-glucosidase activity. We used one-way ANOVA to see the statistical difference among the groups. The significance thresholds were considered at the P < 0.05 level. In comparison with the healthy group, the extract showed a significant antidiabetic effect. The proportion of inhibition increased as the concentrations increased. Previous studies established the antiviral, anti-inflammatory, analgesic, antifungal, antioxidant, antibacterial, hepatoprotective, antiasthmatic, immunomodulatory, and antipyretic effects of Kabasura Kudineer or Choornam. The current findings demonstrated that the Chooranam has good antidiabetic action at a significant concentration. Plant-based products have recently proven to be effective and economical antidiabetic items.

Identification and characterization of conserved noncoding cis-regulatory elements that impact Mecp2 expression and neurological functions.

While changes in MeCP2 dosage cause Rett syndrome (RTT) and MECP2 duplication syndrome (MDS), its transcriptional regulation is poorly understood. Here, we identified six putative noncoding regulatory elements of Mecp2, two of which are conserved in humans. Upon deletion in mice and human iPSC-derived neurons, these elements altered RNA and protein levels in opposite directions and resulted in a subset of RTT- and MDS-like behavioral deficits in mice. Our discovery provides insight into transcriptional regulation of Mecp2/MECP2 and highlights genomic sites that could serve as diagnostic and therapeutic targets in RTT or MDS.

Influence of genetic copy number variants of the human GLUT3 glucose transporter gene SLC2A3 on protein expression, glycolysis and rheumatoid arthritis risk: A genetic replication study.

OBJECTIVES: The gene encoding glucose transporter 3 (GLUT3, SLC2A3) is present in the human population at variable copy number. An overt disease phenotype of SLC2A3 copy number variants has not been reported; however, deletion of SLC2A3 has been previously reported to protect carriers from rheumatoid arthritis, implicating GLUT3 as a therapeutic target in rheumatoid arthritis. Here we aim to perform functional analysis of GLUT3 copy number variants in immune cells, and test the reported protective association of the GLUT3 copy number variants for rheumatoid arthritis in a genetic replication study. METHODS: Cells from genotyped healthy controls were analyzed for SLC2A3/GLUT3 expression and glycolysis capacity. We genotyped the SLC2A3 copy number variant in four independent cohorts of rheumatoid arthritis and controls and one cohort of multiple sclerosis and controls. RESULTS: Heterozygous deletion of SLC2A3 correlates directly with expression levels of GLUT3 and influences glycolysis rates in the human immune system. The frequency of the SLC2A3 copy number variant is not different between rheumatoid arthritis, multiple sclerosis and control groups. CONCLUSIONS: Despite a robust SLC2A3 gene copy number dependent phenotype, our study of large groups of rheumatoid arthritis cases and controls provides no evidence for rheumatoid arthritis disease protection in deletion carriers. These data emphasize the importance of well powered replication studies to confirm or refute genetic associations, particularly for relatively rare variants.

Transmitter release site organization can predict synaptic function at the neuromuscular junction.

We have investigated the impact of transmitter release site (active zone; AZ) structure on synaptic function by physically rearranging the individual AZ elements in a previously published frog neuromuscular junction (NMJ) AZ model into the organization observed in a mouse NMJ AZ. We have used this strategy, purposefully without changing the properties of AZ elements between frog and mouse models (even though there are undoubtedly differences between frog and mouse AZ elements in vivo), to directly test how structure influences function at the level of an AZ. Despite a similarly ordered ion channel array substructure within both frog and mouse AZs, frog AZs are much longer and position docked vesicles in a different location relative to AZ ion channels. Physiologically, frog AZs have a lower probability of transmitter release compared with mouse AZs, and frog NMJs facilitate strongly during short stimulus trains in contrast with mouse NMJs that depress slightly. Using our computer modeling approach, we found that a simple rearrangement of the AZ building blocks of the frog model into a mouse AZ organization could recapitulate the physiological differences between these two synapses. These results highlight the importance of simple AZ protein organization to synaptic function. NEW & NOTEWORTHY A simple rearrangement of the basic building blocks in the frog neuromuscular junction model into a mouse transmitter release site configuration predicted the major physiological differences between these two synapses, suggesting that transmitter release site structure and organization is a strong predictor of function.

On-field management and return-to-play in sports-related concussion in children: Are children managed appropriately?

OBJECTIVES: On-field management and return-to-play guidelines aim to ensure the identification and appropriate management of the concussed athlete. Compliance with current guidelines in many settings is unknown. We assessed whether key components of current concussion guidelines are being followed in child athletes. DESIGN: Prospective observational study. METHODS: Data were collected from children (5-18 years) presenting to a paediatric emergency department with sport-related concussion via researcher-administered surveys in the emergency department and during a follow up phone call. On hospital discharge all patients received a return to sports fact sheet based on the International Concussion in Sports Group. RESULTS: Ninety-three had sustained a concussion (mean age 12.7 (±0.27) years, 83% male). Sports played included Australian Football (47%), soccer (12%), rugby (9%) basketball (8%), other (25%). 82% participated in organised sports. Concussive signs or symptoms included loss of consciousness (41%), disorientation (36%), vomiting (23%), amnesia (30%), headache (60%). For concussive injury in organised sports (n=76), overall 42% were not managed according to recommended guidelines: 19% were not immediately removed from play, 29% were allowed to return to play on the same day and 27% were not assessed by qualified personnel. 93% of parents and 96% of patients were unaware of concussion or return-to-play guidelines from their organisations. Overall, 72% were compliant with provided return-to-play guidelines. CONCLUSIONS: Many children with sports related-concussion are not formally assessed on-field and continue to play. On-field concussion management and return to play practices are often suboptimal. Awareness and education of coaches, teachers, parents and children need to be improved.

Effect of Titanium dioxide nanoparticles on the flexural strength of polymethylmethacrylate: an in vitro study.

CONTEXT: To improve the flexural strength of polymethylmethacrylate (PMMA). AIM: To evaluate whether the incorporation of titanium dioxide nanoparticles in polymethylmethacrylate (PMMA) increases the flexural strength and to compare the different concentrations of titanium dioxide nanoparticles and its relation to flexural strength. SETTINGS AND DESIGN: Study was conducted in Sri Ramachandra University utilizing 40 specimens manufactured from clear heat polymerizing acrylic resin. MATERIALS AND METHODS: Forty specimens of clear heat polymerizing acrylic resin of dimensions 65 Χ 10 Χ 3 mm as per ISO 1,567 standardization were fabricated and were grouped into A (CONTROL) with no titanium dioxide (TiO2) nanoparticles, B with 0.5 gms of TiO 2 nanoparticles, C with 1 gm of TiO 2 nanoparticles and D with 2.5 gms of TiO 2 nanoparticles added.The concentrations of titanium dioxide in each group were 1 wt%, 2 wt% and 5 wt%. Universal testing machine INSTRON was used to load at the center of the specimen with a cross head speed of 1.50 mm/min and a span length of 40.00 mm. STATISTICAL ANALYSIS USED: ANOVA and multiple comparisons are carried out using the independent t-test. RESULTS: The ANOVA result shows that there is a significant difference between the groups with respect to the mean flexural strength. Highest mean flexural strength is observed in Group D, while the lowest is seen in Group A. Independent t-test revealed that there was a statistical significance between Group A and Group D (0.041) and between Group B and Group D (0.028). CONCLUSIONS: The results concluded that polymethylmethacrylate reinforced with different concentrations of titanium dioxide nanoparticles showed superior flexural strength than those of normal PMMA.

Stemness of the CT-2A Immunocompetent Mouse Brain Tumor Model: Characterization In Vitro.

Evidence has pointed to brain tumor stem cells (BTSC) as culprits behind human high-grade glioma (hHGG) resistance to standard therapy. Pre-clinical rodent models are the mainstay for testing of new therapeutic strategies. The typical model involves the intracranial injection of human glioma cells into immunocompromised hosts, hindering the evaluation of tumor-host responses and resulting in non-infiltrative tumors. The CT-2A model is an immunocompetent mouse model with potential to overcome these disadvantages. In this study, we confirmed the highly infiltrative nature of intracranial CT-2A tumors and optimized reproducible injection parameters. We then generated neurospheres and established, for the first time, the stemness of this model. CT-2A expression of the BTSC marker, CD133, increased from 2% in monolayer cells to 31% in fully-formed neurospheres. Investigation of three stem cell markers (Oct4, Nanog and Nestin) revealed a distinct stemness signature with monolayer cells expressing Oct4 and Nestin (no Nanog), and neurospheres expressing all three. Additionally, CT-2A cells were more proliferative and invasive than U87 cells, while CT-2A neurospheres were significantly more proliferative and invasive than either monolayer cells in vitro. Taken together, our results show that this model is a valuable tool for pre-clinical testing of novel therapeutics against hHGG and also affords the opportunity for investigation of BTSC in an immunocompetent setting.

Efficient differentiation of human embryonic and induced pluripotent stem cells into functional astrocytes.

Human high-grade gliomas (hHGG) remain a therapeutic challenge in neuro-oncology despite current multimodality treatments. We recently demonstrated that murine embryonic stem cell (mESC)-derived astrocytes conditionally expressing proapoptotic genes can successfully be used to induce apoptosis and tumor shrinkage of hHGG tumor in vitro and in an in vivo mouse model. The first step in the translation of these results to the clinical settings, however, requires availability of human embryonic stem cells (hESC)- and/or induced pluripotent cell (hiPSC)-derived astrocytes engineered to express proapoptotic genes. The potential for directed differentiation of hESCs and hiPSCs to functional postmitotic astrocytes is not fully characterized. In this study, we show that once specified to neuro-epithelial lineage, hiPSC could be differentiated to astrocytes with a similar efficiency as hESC. However, our analyses of 2 hESC and 2 hiPSC cell lines showed some variability in differentiation potential into astrocytic lineages. Both the hESC- and hiPSC-derived astrocytes appeared to follow the functional properties of mESC-derived astrocytes, namely, migration and tropism for hHGG. This work provides evidence that hESC- and hiPSC-derived cells are able to generate functionally active astrocytes. These results demonstrate the feasibility of using iPSC-derived astrocytes, a new potential source for therapeutic use for brain tumors and other neurological diseases.

Is there a common upstream link for autophagic and apoptotic cell death in human high-grade gliomas?

The prognosis of patients with human high-grade gliomas (HGGs) remains dismal despite major advances in their management, due mainly to the high resistance of these infiltrative tumor cells to programmed cell death (PCD). Most therapeutic strategies for HGGs are aimed to maximize PCD type I, apoptosis or type II, autophagy. These are predominantly distinctive processes, but many studies suggest a cross-talk between the two. A better understanding of the link between PCD types I and II might allow development of more effective therapies for HGGs. In this study, we examined whether there is a common upstream signaling event responsible for both apoptotic and autophagic PCD using 3 chemotherapeutic agents in human HGG cells. Our study shows that each agent caused a significant decrease in cell viability in each of the HGG cell lines tested. The increase rate of apoptosis and autophagy varied among cell lines and chemotherapeutic agents used. Increased expression of cytidine-cytidine-adenosine-adenosine-thymidine (C)/enhancer binding protein (EBP) homologous transcription factor C/EBP homologous protein (CHOP)/growth arrest and DNA damage-inducible gene 153 (GADD153) was documented after use of either pro-autophagic or pro-apoptotic agents. The involvement of CHOP/GADD153 in both type I and type II PCD was confirmed by overexpression and gene-silencing studies. Gene silencing by small-interfering RNA-mediated CHOP/GADD153 resulted in increased cell viability, decreased upregulation of microtubule-associated protein light-chain 3' type II (LC3II) and cleaved caspase-3, and inhibition of apoptosis and autophagy. Exogenous expression of CHOP/GADD153 triggered apoptosis and autophagy in the absence of other stimuli. The clinical significance of these findings was supported by the evidence that celecoxib, a nonsteroidal anti-inflammatory drug known to induce GADD153-mediated apoptosis, strongly increases both type I and type II PCD in HGG cells when combined with another inducer of GADD153. These data suggest that CHOP/GADD153 should be investigated as a novel targetable signaling step to improve therapies for HGGs.

Efficacy of a probiotic and chlorhexidine mouth rinses: a short-term clinical study.

INTRODUCTION: Probiotic technology represents a breakthrough approach to maintaining oral health by utilizing natural beneficial bacteria commonly found in healthy mouths to provide a natural defense against those bacteria thought to be harmful to teeth and gums. However, data are still sparse on the probiotic action in the oral cavity. The review article on probiotics in children published by Twetman and Stecksen- Blicks in 2008 showed only one study of dental interest on probiotics in children. AIM AND OBJECTIVES: The present study evaluated clinically the efficacy of a probiotic and chlorhexidine mouth rinses on plaque and gingival accumulation in children. The trial design is a double-blind parallel group, 14 days comparative study between a probiotic mouth rinse and a chlorhexidine mouth rinse, which included 45 healthy children in the age group of 6-8 years. RESULTS: The Probiotic and Chlorhexidine groups had less plaque accumulations compared with the Control group at the end of 14 years (P < 0.001 and P < 0.001, respectively). But, unlike the plaque score, there was a significant difference in the Gingival Index between the Probiotic and the Chlorhexidine groups (P = 0.009), Probiotic group being better than the Chlorhexidine group (mean = 0.2300 and 0.6805, respectively). CONCLUSION: The Probiotic mouth rinse was found effective in reducing plaque accumulation and gingival inflammation. Therefore, probiotic mouth rinse obviously has a potential therapeutic value and further long-term study is recommended to determine its efficacy.