Genetic Risk Factors for Early-Onset Merkel Cell Carcinoma.

Importance: Merkel cell carcinoma (MCC) is a rare, aggressive neuroendocrine skin cancer. Of the patients who develop MCC annually, only 4% are younger than 50 years. Objective: To identify genetic risk factors for early-onset MCC via genomic sequencing. Design, Setting, and Participants: The study represents a multicenter collaboration between the National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS), the National Institute of Allergy and Infectious Diseases (NIAID), and the University of Washington. Participants with early-onset and later-onset MCC were prospectively enrolled in an institutional review board-approved study at the University of Washington between January 2003 and May 2019. Unrelated controls were enrolled in the NIAID Centralized Sequencing Program (CSP) between September 2017 and September 2021. Analysis was performed from September 2021 and March 2023. Early-onset MCC was defined as disease occurrence in individuals younger than 50 years. Later-onset MCC was defined as disease occurrence at age 50 years or older. Unrelated controls were evaluated by the NIAID CSP for reasons other than familial cancer syndromes, including immunological, neurological, and psychiatric disorders. Results: This case-control analysis included 1012 participants: 37 with early-onset MCC, 45 with later-onset MCC, and 930 unrelated controls. Among 37 patients with early-onset MCC, 7 (19%) had well-described variants in genes associated with cancer predisposition. Six patients had variants associated with hereditary cancer syndromes (ATM = 2, BRCA1 = 2, BRCA2 = 1, and TP53 = 1) and 1 patient had a variant associated with immunodeficiency and lymphoma (MAGT1). Compared with 930 unrelated controls, the early-onset MCC cohort was significantly enriched for cancer-predisposing pathogenic or likely pathogenic variants in these 5 genes (odds ratio, 30.35; 95% CI, 8.89-106.30; P < .001). No germline disease variants in these genes were identified in 45 patients with later-onset MCC. Additional variants in DNA repair genes were also identified among patients with MCC. Conclusions and Relevance: Because variants in certain DNA repair and cancer predisposition genes are associated with early-onset MCC, genetic counseling and testing should be considered for patients presenting at younger than 50 years.

Assessment of left ventricular function in patients with type 2 diabetes mellitus by non-invasive myocardial work.

Background: Diabetes mellitus (DM) is a chronic disease that poses a serious risk of cardiovascular diseases. Therefore, early detection of impaired cardiac function with non-invasive myocardial imaging is critical for improving the prognosis of patients with DM. Purpose: This study aimed to assess the left ventricular (LV) function in patients with type 2 diabetes mellitus (T2DM) by non-invasive myocardial work technique. Materials and methods: In all, 67 patients with T2DM and 28 healthy controls were included and divided into a DM group and a control group. Two-dimensional dynamic images of apical three-chamber view, apical two-chamber view, and apical four-chamber view were collected from all subjects, consisting of at least three cardiac cycles. LV myocardial strain parameters, including global longitudinal strain (GLS) and peak strain dispersion (PSD), as well as myocardial work parameters, including global constructive work (GCW), global wasted work (GWW), global work index (GWI), and global work efficiency (GWE), were obtained and analyzed. Results: A total of 15 subjects were randomly selected to assess intra-observer and inter-observer consistency of myocardial work parameters and strain parameters, which showed excellent results (intra-class correlation coefficients: 0.856 - 0.983, P<0.001). Compared with the control group, the DM group showed significantly higher PSD (37.59 ± 17.18 ms vs. 27.72 ± 13.52 ms, P<0.05) and GWW (63.98 ± 43.63 mmHg% vs. 39.28 ± 25.67 mmHg%, P<0.05), and lower GWE (96.38 ± 2.02% vs. 97.72 ± 0.98%, P<0.001). Furthermore, the PSD was positively correlated with GWW (r = 0.565, P<0.001) and negatively correlated with GWE (r = -0.569, P<0.001). Conclusion: Uncoordinated LV myocardial strain, higher GWW, and lower GWE in patients with T2DM may serve as indicators for the early assessment of cardiac impairment in T2DM.

Chromosomal microarray analysis supplements exome sequencing to diagnose children with suspected inborn errors of immunity.

Purpose: Though copy number variants (CNVs) have been suggested to play a significant role in inborn errors of immunity (IEI), the precise nature of this role remains largely unexplored. We sought to determine the diagnostic contribution of CNVs using genome-wide chromosomal microarray analysis (CMA) in children with IEI. Methods: We performed exome sequencing (ES) and CMA for 332 unrelated pediatric probands referred for evaluation of IEI. The analysis included primary, secondary, and incidental findings. Results: Of the 332 probands, 134 (40.4%) received molecular diagnoses. Of these, 116/134 (86.6%) were diagnosed by ES alone. An additional 15/134 (11.2%) were diagnosed by CMA alone, including two likely de novo changes. Three (2.2%) participants had diagnostic molecular findings from both ES and CMA, including two compound heterozygotes and one participant with two distinct diagnoses. Half of the participants with CMA contribution to diagnosis had CNVs in at least one non-immune gene, highlighting the clinical complexity of these cases. Overall, CMA contributed to 18/134 diagnoses (13.4%), increasing the overall diagnostic yield by 15.5% beyond ES alone. Conclusion: Pairing ES and CMA can provide a comprehensive evaluation to clarify the complex factors that contribute to both immune and non-immune phenotypes. Such a combined approach to genetic testing helps untangle complex phenotypes, not only by clarifying the differential diagnosis, but in some cases by identifying multiple diagnoses contributing to the overall clinical presentation.

Clinical exome sequencing of 1000 families with complex immune phenotypes: Toward comprehensive genomic evaluations.

BACKGROUND: Prospective genetic evaluation of patients at this referral research hospital presents clinical research challenges. OBJECTIVES: This study sought not only a single-gene explanation for participants' immune-related presentations, but viewed each participant holistically, with the potential to have multiple genetic contributions to their immune phenotype and other heritable comorbidities relevant to their presentation and health. METHODS: This study developed a program integrating exome sequencing, chromosomal microarray, phenotyping, results return with genetic counseling, and reanalysis in 1505 individuals from 1000 families with suspected or known inborn errors of immunity. RESULTS: Probands were 50.8% female, 71.5% were ≥18 years, and had diverse immune presentations. Overall, 327 of 1000 probands (32.7%) received 361 molecular diagnoses. These included 17 probands with diagnostic copy number variants, 32 probands with secondary findings, and 31 probands with multiple molecular diagnoses. Reanalysis added 22 molecular diagnoses, predominantly due to new disease-gene associations (9 of 22, 40.9%). One-quarter of the molecular diagnoses (92 of 361) did not involve immune-associated genes. Molecular diagnosis was correlated with younger age, male sex, and a higher number of organ systems involved. This program also facilitated the discovery of new gene-disease associations such as SASH3-related immunodeficiency. A review of treatment options and ClinGen actionability curations suggest that at least 251 of 361 of these molecular diagnoses (69.5%) could translate into ≥1 management option. CONCLUSIONS: This program contributes to our understanding of the diagnostic and clinical utility whole exome analysis on a large scale.

Clinical Outcomes of Aberration-Free All Surface Laser Ablation (ASLA) vs. Aberration-Free ASLA Assisted by Smart Pulse Technology in High Myopia: A One-Year Follow-Up Study.

PURPOSE: To compare the clinical outcomes of aberration-free all surface laser ablation (ASLA) with and without the use of smart pulse technology (SPT) in high myopia. METHODS: This study retrospectively analyzed 138 eyes (138 patients, only the right eye was selected) treated for high myopia (spherical equivalent ≥-6.00 diopters) using aberration-free ASLA (non-SPT group; 85 eyes) and aberration-free ASLA assisted by SPT (SPT group; 53 eyes). Examinations such as visual acuity, refraction, and haze were performed before the 12-month follow-up. Corneal epithelial healing time was assessed in the first postoperative day. Visual acuity and refraction examination were performed at 7 days and 1, 3, 6, and 12 months postoperatively. Corneal haze was evaluated in 1, 3, 6, and 12 months. Safety, efficacy, and corneal wavefront aberrations were assessed 12 months after the treatment. RESULTS: At 12 months postoperatively, 60% versus 40% of eyes achieved 20/16 Snellen lines or better, and 92% versus 82% of eyes achieved 20/20 Snellen lines or better visual acuity in the SPT and the non-SPT groups, respectively. The average postoperative epithelial healing time was 3.75 ± 1.00 days in the SPT group and 3.73 ± 1.30 days in the non-SPT group (P ≥ 0.05). The safety and the efficacy index of the SPT group were better than those of the non-SPT group in the follow-ups. The attempted spherical equivalent before the surgery and the achieved spherical equivalent at 12 months were comparable between the two groups. Regarding the aberrations, the results of Coma 90° in the SPT group were better than those in the non-SPT group (P ≤ 0.05), but the increase of RMS HOAs (root mean square higher order aberrations), Coma 0°, and spherical aberration postoperatively had no statistical difference between the two groups (P ≥ 0.05). Conclusions: Both aberration-free ASLA with and without SPT showed favorable safety, effectiveness, and predictability within 12 months for high myopia. And, ASLA using SPT might have potential advantages in the long-term visual quality.

The molecular landscape of neural differentiation in the developing Drosophila brain revealed by targeted scRNA-seq and multi-informatic analysis.

The Drosophila type II neuroblast lineages present an attractive model to investigate the neurogenesis and differentiation process as they adapt to a process similar to that in the human outer subventricular zone. We perform targeted single-cell mRNA sequencing in third instar larval brains to study this process of the type II NB lineage. Combining prior knowledge, in silico analyses, and in situ validation, our multi-informatic investigation describes the molecular landscape from a single developmental snapshot. 17 markers are identified to differentiate distinct maturation stages. 30 markers are identified to specify the stem cell origin and/or cell division numbers of INPs, and at least 12 neuronal subtypes are identified. To foster future discoveries, we provide annotated tables of pairwise gene-gene correlation in single cells and MiCV, a web tool for interactively analyzing scRNA-seq datasets. Taken together, these resources advance our understanding of the neural differentiation process at the molecular level.

Pollution Distribution of Potentially Toxic Elements in a Karstic River Affected by Manganese Mining in Changyang, Western Hubei, Central China.

This study investigated the distribution, pollution level and potential ecological risk of potentially toxic elements (PTEs) from manganese mining in a karstic Danshui River, in Changyang, Western Hubei, Central China. River water and sediments were collected for seven PTEs measurement (As, Cd, Cr, Cu, Mn, Pb and Zn), as well as pH and Eh of the river water were measured. Results showed that the major pollutant was Mn, the river water environment was mainly acidic and oxidizing (288 < Eh, pH < 6.3), and the pollution distribution of Mn in the study area was dominated by the combination of natural processes and anthropogenic activities. In the river water, according to the contamination factor (CF) and pollution load index (IPL) results, Mn was considered the main pollutant. There was low As and Pb pollution downstream as well as Cu pollution upstream. Upstream and downstream areas were the main polluted river sections of the river water samples collected. In river sediments, based on the results of the geo-accumulation index (Igeo) and potential ecological risk index (IPER), it was determined that there was only considerable Mn pollution. The IPER of the PTEs from the river sediments was at acceptable levels, only Mn upstream performed at a moderate ecological risk level. According to Pearson correlation and principal component analysis, Mn originated from manganese mining activities, Cd, Cr and Zn were of natural origin, and Cu may have come from both mining and natural origin, whereas Pb and As were mainly related to the daily activities. Consequently, elemental speciation, mining activities and the distribution of water conservancy facilities were the main impacts of PET pollution distribution in this river.

Co-overexpression of AtSHMT1 and AtFDH induces sugar synthesis and enhances the role of original pathways during formaldehyde metabolism in tobacco.

Serine hydroxymethyltransferase 1 (SHMT1) is a key enzyme in the photorespiration pathway in higher plants. Our previous study showed that AtSHMT1 controls the assimilation of HCHO to sugars in Arabidopsis. The expression of SHMT1 was induced in Arabidopsis but was inhibited in tobacco under HCHO stress. To investigate whether the function of AtSHMT1 in the HCHO assimilation could be exerted in tobacco, AtSHMT1 was overexpressed alone (S5) or co-overexpressed (SF6) with Arabidopsis formate dehydrogenase (AtFDH) in leaves using a light-inducible promoter in this study. 13C NMR analyses showed that the 13C-metabolic flux from H13CHO was introduced to sugar synthesis in SF6 leaves but not in S5 leaves. The increase in the production of metabolites via the original pathways was particularly greater in SF6 leaves than in S5 leaves, suggesting that co-overexpression of AtSHMT1 and AtFDH is more effective than overexpression of AtSHMT1 alone in the enhancement of HCHO metabolism in tobacco leaves. Consequently, the increase in HCHO uptake and resistance was greater in SF6 leaves than in S5 leaves. The mechanism underlying the role of overexpressed AtSHMT1 and AtFDH was discussed based on changes in photosynthetic parameters, chlorophyll content, antioxidant enzyme activity and the oxidative level in leaves.

[Seasonal Distribution Characteristics and Health Risk Assessment of Heavy Metals in Surface Water of Qingjiang River].

In order to assess the health risks of heavy metals in surface water of Qingjiang River, surface water samples were taken at designed cross-sections of the river and analyzed for Cr, Cu, Zn, Pb, Cd, As, and Mn. Health risks from these heavy metals for adults and children in wet and dry seasons were compared by water environmental health risk assessment model of the USEPA. It found that the main excessive element is Mn, concentrating in the Danshui, Yantouxi, and Pingluoxi, the slightly excessive element is As, the concentration of Mn was above national standard, and it mainly distributed in Danshui, Yantouxi, and Pingluoxi, As was slightiy over the standard, and it concentrated in Wujiahe, The content of heavy metals during wet season were all higher than those during dry season. Cr, Cu, Zn, and Cd are mainly originated from the nature, Pb and As are separately mainly originated from traffic and agriculture, Mn originated from mining mainly in the downstream, while it has natural source from upper to middle. The health risks of heavy metals in surface water to adults and children in wet season are higher than those in dry season. The main health risk area was the midstream. As was the highest health risk element and children were the most preventive group. Specially, people in towns who drink the water from midstream should pay more attention.

Formaldehyde assimilation through coordination of the glyoxylate pathway and the tricarboxylic acid cycle in broad bean roots.

Formaldehyde (HCHO) assimilation in broad bean (Vicia faba L. cv. YD) roots was investigated using 13C-labeled HCHO followed by 13C-NMR analysis. Results revealed that H13CHO was first oxidized to H13COOH in the roots treated with 2 mM H13CHO in a time-dependent manner. Subsequently, a massive signal peak of [2, 4-13C]citrate (Cit) and a signal peak of [2, 3-13C]succinate (Su) were observed in accompany with an enhancement in the signal intensity of [3-13C]Cit. The data suggested that the glyoxylate pathway and the tricarboxylic acid (TCA) cycle functioned simultaneously in the subsequent assimilation of H13COOH. The yield of [2, 4-13C]Cit accounted for more than 80% of the total metabolites. The activity of isocitrate lyase (ICL), a key enzyme in the glyoxylate pathway, was stimulated by HCHO in a dosage-dependent manner. As a result, [2, 4-13C]Cit production was increased significantly in YD roots treated with high concentrations (4 and 6 mM) of H13CHO. Moreover, induction of the ICL activity by methanol resulted in a simultaneous elevation in the production of [2, 4-13C]Cit and [3-13C]Cit in methanol-pretreated roots under 2 mM H13CHO stress. Pretreatment of roots with cyclosporin A, which hinders the transport of 13C-enriched compounds into mitochondria, caused a notable decline in the signal peak and yield of [2, 4-13C]Cit and consequently induced a notable accumulation of [2, 3-13C]Su and an increase in the HCO3- production (generated from H13COOH oxidation) in H13CHO-treated roots. These results suggested that the glyoxylate pathway and the TCA cycle function coordinately in HCHO assimilation in broad bean roots.

Hypoxia-regulated lncRNA CRPAT4 promotes cell migration via regulating AVL9 in clear cell renal cell carcinomas.

INTRODUCTION: Long noncoding RNAs (lncRNAs) are proven to be key regulators in cancer biology. Our screening effort for clear cell renal cell carcinoma (ccRCC) prognosis-associated lncRNAs identified a novel lncRNA, ccRCC prognosis-associated transcript 4 (CRPAT4), as one of the top candidates that was previously uncharacterized. The aim of this study was to verify the clinical significance of CRPAT4 in ccRCC patients and to explore its biological role as well as the underlying mechanisms, in ccRCC cell lines. MATERIALS AND METHODS: Quantitative real-time polymerase chain reaction (PCR) was performed to demonstrate that CRPAT4 was differentially expressed between ccRCC and the normal controls and that high CRPAT4 expression significantly associated with advanced Fuhrman nuclear grades. RESULTS: Kaplan-Meier survival analysis with The Cancer Genome Atlas KIRC RNA sequencing data indicated that high CRPAT4 expression was significantly associated with poor overall survival and progression-free survival. Functional studies indicated that CRPAT4 was an HIF-1α regulated gene, and CRPAT4 knockdown significantly inhibited cell migration and proliferation in the absence of HIF-1α. In addition, a mechanistic study revealed that CRPAT4 could regulate the expression of the migration-associated protein AVL9. CONCLUSION: Collectively, our study first identified CRPAT4 as a hypoxia-regulated lncRNA, acting as an oncogene in ccRCC progression via regulating AVL9 protein, thus expanding our knowledge on the hypoxia pathway in ccRCC biology from a noncoding perspective. Moreover, CRPAT4 has the potential to be a prognostic marker in ccRCC patients.

Overexpression of p42.3 promotes cell proliferation, migration, and invasion in human gastric cancer cells.

As a newly discovered tumor-specific gene, p42.3 is overexpressed in most of human gastric cancers (GC). However, the role of p42.3 in GC progression remains unclear. To assess the role of p42.3 in gastric cancers, immunohistochemistry and western blot were performed to detect the p42.3 expression in human GC tissues and cells. We also investigated the role of p42.3 in GC cell proliferation, migration, and invasion. Our results showed that the p42.3 expression was increased dramatically in human GC tissue and cells. In addition, we found that overexpression of p42.3 promotes GC cell proliferation, migration, and invasion abilities. Furthermore, p42.3 expression suppressed the E-cadherin protein level and promoted the ß-catenin and p-ERK protein level. Taken together, overexpressed p42.3 is correlated with gastric cancer cell proliferation, migration, and invasion, suggesting its use as a biological marker in gastric cancer.

Rap2B promotes proliferation, migration, and invasion of human breast cancer through calcium-related ERK1/2 signaling pathway.

Rap2B, a member of GTP-binding proteins, is widely upregulated in many types of tumors and promotes migration and invasion of human suprarenal epithelioma. However, the function of Rap2B in breast cancer is unknown. Expression of Rap2B was examined in breast cancer cell lines and human normal breast cell line using Western blot analysis. Using the CCK-8 cell proliferation assay, cell cycle analysis, and transwell migration assay, we also elucidated the role of Rap2B in breast cancer cell proliferation, migration, and invasion. Results showed that the expression of Rap2B is higher in tumor cells than in normal cells. Flow cytometry and Western blot analysis revealed that Rap2B elevates the intracellular calcium level and further promotes extracellular signal-related kinase (ERK) 1/2 phosphorylation. By contrast, calcium chelator BAPTM/AM and MEK inhibitor (U0126) can reverse Rap2B-induced ERK1/2 phosphorylation. Furthermore, Rap2B knockdown inhibits cell proliferation, migration, and invasion abilities via calcium related-ERK1/2 signaling. In addition, overexpression of Rap2B promotes cell proliferation, migration and invasion abilities, which could be neutralized by BAPTM/AM and U0126. Taken together, these findings shed light on Rap2B as a therapeutic target for breast cancer.

p21-Activated kinase 5 affects cisplatin-induced apoptosis and proliferation in hepatocellular carcinoma cells.

p21-Activated kinase 5 (PAK5) is the last identified member of the PAK family. The PAKs are highly conserved serine/threonine and effector proteins for Cdc42 and Rac and are essential in regulating cell motility and survival. Previous studies have demonstrated that PAK5 played a pivotal role in apoptosis, proliferation, cancer migration, and invasion. However, the biological function of PAK5 in hepatocellular carcinoma, as well as its underlying mechanism, still remains to be fully elucidated. In the present study, we demonstrated that PAK5 markedly inhibited cisplatin-induced apoptosis and promoted cell proliferation in hepatocellular carcinoma cells. Moreover, our results showed that overexpression of PAK5 contributed to cell cycle regulation. In order to elucidate the underlying mechanism of PAK5 on cisplatin-induced apoptosis and cell cycle regulation, we also examined the protein expressions of chk2 and p-chk2. In summary, our study investigated the role of PAK5 in cisplatin-induced cellular processes and provided evidence of its underlying mechanism.

p42.3: An abductor of cell cycle.

As a newly discovered tumor-associated gene, p42.3 was originally ascertained in gastric cancer cell line BGC823 and has been confirmed as a cell cycle-dependent gene that is overexpressed in many human tumor cell lines and embryonic tissues. p42.3 can regulate the level of relevant cycle-dependent proteins and promote malignant transformation of cells. A variety of cellular functions, including cell proliferation, cell invasion and cell migration, are under control of p42.3. Our review, namely the introduction of the structure of p42.3, underlying activity regulation mechanisms of p42.3 as well as the role p42.3 plays in malignant cellular transformation process, are accompanied by the presentation of potential directions of further researches of cancer prevention and therapy in which p42.3 is inevitable.

[Identification analysis of eukaryotic expression plasmid Rap2a and its effect on the migration of lung cancer cells].

BACKGROUND: Rap2a, a member of the small GTPase superfamily, plays a critical role in regulating the function of integrin and cell adhesion, thereby controlling cell motility and cell/matrix interactions. However, the function of Rap2a in carcinogenesis is still poorly understood. To clone Rap2a cDNA, which belongs to human Ras-related small G protein superfamily, we constructed its eukaryotic expression vector and determined its expression in lung cancer cells. The aim of this study is to explore the role of Rap2a in carcinogenesis. METHODS: The levels of endogenous Rap2a protein in lung cancer cells were measured by Western blot. Total RNA of human osteosarcoma cells U2OS was extracted and reverse-transcribed into cDNA by RT-PCR. Then, Rap2a gene was amplified by PCR and inserted into pcDNA3.1(+). The reconstructed plasmid was identified by restricted enzyme digestion and sequencing. pcDNA3.1(+)-Rap2a was transfected into H1299 and A549 cells, the expression of Rap2a was detected by Western blot. In addition, the migratory abilities of lung cancer cells were evaluated by Transwell assay. Matrix metalloproteinase (MMP)2 enzyme activity was evaluated by gelatin zymography. RESULTS: Rap2a is significantly upregulated in lung cancer cells. The results of enzyme digestion and sequencing showed that the coding sequence of pcDNA3.1(+)-Rap2a was right and was inserted into the vector correctly. The results of Western blot showed that H1299 and A549 cells were transfected successfully. Transwell assay indicated that the ectopic expression of Rap2a promotes lung cancer cells migration. Correspondly, enzyme activity of MMP2 also increased. CONCLUSIONS: Eukaryotic expression plasmid pcDNA3.1(+)-Rap2a was constructed successfully. Rap2a could be expressed in lung cancer cells efficiently and promotes lung cancer cell migration.

p42.3 promotes cell proliferation and invasion in human Renal-Cell Carcinoma.

p42.3 is a tumor-specific gene and found to be over-expressed in many tumor cell lines and primary tumor tissues. It plays a significant role in neoplastic transformation and tumor progression. To date, the association between p42.3 and Renal-Cell Carcinoma (RCC) has not been reported. This study investigated the biological effects and mechanisms of p42.3 in RCC progression. In this study, we found that p42.3 is overexpressed in various kinds of RCC cells, and knockdown of p42.3 dramatically reduced cell proliferation and invasion in vitro. Our studies revealed that overexpression of p42.3 accelerates the epithelial-mesenchymal transition (EMT) progression and induces RCC cells proliferation and invasion. Further studies show that p42.3 may involve in activation of ß-catenin and participate in RCC cell invasion. Combined, these data indicate that p42.3 contributes to promoting RCC cells proliferation and invasion through accelerates the EMT progression and ß-catenin activation.