Multi-modal molecular determinants of clinically relevant osteoporosis subtypes.

Due to a rapidly aging global population, osteoporosis and the associated risk of bone fractures have become a wide-spread public health problem. However, osteoporosis is very heterogeneous, and the existing standard diagnostic measure is not sufficient to accurately identify all patients at risk of osteoporotic fractures and to guide therapy. Here, we constructed the first prospective multi-omics atlas of the largest osteoporosis cohort to date (longitudinal data from 366 participants at three time points), and also implemented an explainable data-intensive analysis framework (DLSF: Deep Latent Space Fusion) for an omnigenic model based on a multi-modal approach that can capture the multi-modal molecular signatures (M3S) as explicit functional representations of hidden genotypes. Accordingly, through DLSF, we identified two subtypes of the osteoporosis population in Chinese individuals with corresponding molecular phenotypes, i.e., clinical intervention relevant subtypes (CISs), in which bone mineral density benefits response to calcium supplements in 2-year follow-up samples. Many snpGenes associated with these molecular phenotypes reveal diverse candidate biological mechanisms underlying osteoporosis, with xQTL preferences of osteoporosis and its subtypes indicating an omnigenic effect on different biological domains. Finally, these two subtypes were found to have different relevance to prior fracture and different fracture risk according to 4-year follow-up data. Thus, in clinical application, M3S could help us further develop improved diagnostic and treatment strategies for osteoporosis and identify a new composite index for fracture prediction, which were remarkably validated in an independent cohort (166 participants).

Advances in the metabolic engineering of Saccharomyces cerevisiae and Yarrowia lipolytica for the production of ß-carotene.

ß-Carotene is one kind of the most important carotenoids. The major functions of ß-carotene include the antioxidant and anti-cardiovascular properties, which make it a growing market. Recently, the use of metabolic engineering to construct microbial cell factories to synthesize ß-carotene has become the latest model for its industrial production. Among these cell factories, yeasts including Saccharomyces cerevisiae and Yarrowia lipolytica have attracted the most attention because of the: security, mature genetic manipulation tools, high flux toward carotenoids using the native mevalonate pathway and robustness for large-scale fermentation. In this review, the latest strategies for ß-carotene biosynthesis, including protein engineering, promoters engineering and morphological engineering are summarized in detail. Finally, perspectives for future engineering approaches are proposed to improve ß-carotene production.

High-Level Production of Patchoulol in Yarrowia lipolytica via Systematic Engineering Strategies.

Patchoulol is an important sesquiterpene alcohol with a strong and lasting odor, which has led to prominent applications in perfumes and cosmetics. In this study, systematic metabolic engineering strategies were adopted to create an efficient yeast cell factory for patchoulol overproduction. First, a baseline strain was constructed by selecting a highly active patchoulol synthase. Subsequently, the mevalonate precursor pool was expanded to boost patchoulol synthesis. Moreover, a method for downregulating squalene synthesis based on Cu2+-repressible promoter was optimized, which significantly improved the patchoulol titer by 100.9% to 124 mg/L. In addition, a protein fusion strategy resulted in a final titer of 235 mg/L in shake flasks. Finally, 2.864 g/L patchoulol could be produced in a 5 L bioreactor, representing a remarkable 1684-fold increase compared to the baseline strain. To our knowledge, this is the highest patchoulol titer reported so far.

High-yield α-humulene production in Yarrowia lipolytica from waste cooking oil based on transcriptome analysis and metabolic engineering.

BACKGROUND: α-Humulene is an important biologically active sesquiterpene, whose heterologous production in microorganisms is a promising alternative biotechnological process to plant extraction and chemical synthesis. In addition, the reduction of production expenses is also an extremely critical factor in the sustainable and industrial production of α-humulene. In order to meet the requirements of industrialization, finding renewable substitute feedstocks such as low cost or waste substrates for terpenoids production remains an area of active research. RESULTS: In this study, we investigated the feasibility of peroxisome-engineering strain to utilize waste cooking oil (WCO) for high production of α-humulene while reducing the cost. Subsequently, transcriptome analysis revealed differences in gene expression levels with different carbon sources. The results showed that single or combination regulations of target genes identified by transcriptome were effective to enhance the α-humulene titer. Finally, the engineered strain could produce 5.9 g/L α-humulene in a 5-L bioreactor. CONCLUSION: To the best of our knowledge, this is the first report that converted WCO to α-humulene in peroxisome-engineering strain. These findings provide valuable insights into the high-level production of α-humulene in Y. lipolytica and its utilization in WCO bioconversion.

Recent Development of Advanced Biotechnology in the Oleaginous Fungi for Arachidonic Acid Production.

Arachidonic acid is an essential ω-6 polyunsaturated fatty acid, which plays a significant role in cardiovascular health and neurological development, leading to its wide use in the food and pharmaceutical industries. Traditionally, ARA is obtained from deep-sea fish oil. However, this source is limited by season and is depleting the already threatened global fish stocks. With the rapid development of synthetic biology in recent years, oleaginous fungi have gradually attracted increasing attention as promising microbial sources for large-scale ARA production. Numerous advanced technologies including metabolic engineering, dynamic regulation of fermentation conditions, and multiomics analysis were successfully adapted to increase ARA synthesis. This review summarizes recent advances in the bioengineering of oleaginous fungi for ARA production. Finally, perspectives for future engineering approaches are proposed to further improve the titer yield and productivity of ARA.

[Characteristics and Health Risk Assessment of Trace Elements in Atmospheric PM1 During Autumn and Winter in Qingdao].

From November 1,2018 to January 31,2019 (OP2018-2019) and from November 1,2019 to January 20, 2020 (OP2019-2020), PM1 measurement was conducted daily for two consecutive years. The concentration of trace elements in the atmospheric PM1 in Qingdao in autumn and winter was analyzed. The observation period was divided into four air quality levels (Level Ⅰ, Level Ⅱ, Level Ⅲ, and Level Ⅳ), and the characteristics and sources of the concentration of trace elements in PM1 were analyzed. The non-carcinogenic risks (Zn, Pb, Mn, Cu, and V) and carcinogenic risks (As, Cr, Ni, Cd, and Co) of different people with different air quality levels were evaluated. The results showed that the changes in total metal element concentrations were associated with changes in Ca, K, and Al concentrations at different air quality classes during OP2019-2020 compared to those during OP2018-2019 and were more influenced by dust and biomass combustion sources. Compared with that during OP2018-2019, the V concentration in different air quality levels (Level Ⅰ, Level Ⅱ, Level Ⅲ, and Level Ⅳ) during OP2019-2020 decreased by 19.0%, 60.5%, 82.7%, and 77.5%, respectively. This was presumed to be related to the implementation of the Domestic Emission Control Area (DECA) policy for ships, which led to the significant reduction in V concentration due to the change in fuel quality of ships in the waters around Qingdao. The results of the enrichment factor, the ratio method, and the backward trajectory of airflow further indicated that the changes in V concentrations were mainly influenced by the DECA policy. However, after the implementation of the DECA, the V/Ni value as a limit for judging the influence of ship sources in the area required further exploration. The health risk assessment results showed that the risk factor of Mn ranged from 0.07 to 1.22 during OP2018-2019 and OP2019-2020. It was recommended to strengthen the management and control of Mn-containing pollution sources. The lifetime carcinogenic risk (ILCR) value of As and Cd under different air qualities during OP2018-2019 and OP2019-2020 was lower than 10-4 but higher than 10-6, indicating that there was a carcinogenic probability, although it was still at an acceptable level. During OP2018-2019, when the air quality was Ⅳ, the ILCR value of Cr was higher than 10-4, and there was a risk of cancer.

Establishment of relapse risk model and multivariate logistic regression analysis on risk factors of relapse in children with primary nephrotic syndrome.

This study aimed to investigate relapse risk factors in children with primary nephrotic syndrome (PNS) for prevention and early intervention via logistic regression. One hundred thirty-seven children with PNS were enrolled in this study. Clinical variables were analyzed by single-factor and multiple regression analysis to establish the regression equation. The predictive ability of the regression equation was investigated by the receiver operating characteristic curve (ROC). Files of 17 patients were lost, and 120 patients were enrolled finally in the study, among whom 55 cases (45.8%) had frequently relapsed. Single-factor analysis and multiple regression analysis revealed that concurrent infection on first onset, irregular glucocorticoid therapy, severe hypoalbuminemia, and persistent severe hyperlipidemia were the significant risk factors for frequent relapse on PNS (P < .05), among which infection remained to be the main inductive factor. Among the 4 indicators, serum albumin had the best diagnostic efficacy based on the area under the ROC curve (0.933), sensitivity (89.09%), and specificity (81.54%). The area under curve, sensitivity, and specificity for the combined diagnostic model of the 4 indices were 97.8%, 98.18%, and 90.77%, respectively, which had good predictive power for the relapse of patients. Concurrent infection, irregular glucocorticoid therapy, severe hypoalbuminemia, and persistent severe hyperlipemia were all the risk factors for PNS relapse. The established logistic regression model based on these factors above is reliable for predicting frequent PNS relapse. Much attention should be paid to these critical factors, and early intervention should be taken to reduce the incidence of relapse.

Harnessing Yarrowia lipolytica Peroxisomes as a Subcellular Factory for α-Humulene Overproduction.

The sesquiterpene α-humulene has been shown to have anti-inflammatory and anticancer activities, which has led to its vast application potential in medicine. However, α-humulene production methods including phytoextraction and chemical synthesis currently were limited to low yield, high costs, and expensive catalysts, which cannot meet the increasing market demand. In this study, Yarrowia lipolytica was developed as a robust cell factory for α-humulene production. The peroxisome in Y. lipolytica was first engineered to boost the synthesis of the sesquiterpene α-humulene. By compartmentalization of the α-humulene biosynthesis pathway, improving ATP and acetyl-CoA supply, and optimizing the gene copy numbers of rate-limiting enzymes, the engineered strain GQ2012 could produce 3.2 g/L α-humulene in a 5 L bioreactor, the highest α-humulene titer reported so far. Our study provides a valuable reference for highly sustainable production of terpenoids by peroxisome engineering in Y. lipolytica.

YALIcloneNHEJ: An Efficient Modular Cloning Toolkit for NHEJ Integration of Multigene Pathway and Terpenoid Production in Yarrowia lipolytica.

Non-homologous end-joining (NHEJ)-mediated random integration in Yarrowia lipolytica has been demonstrated to be an effective strategy for screening hyperproducer strains. However, there was no multigene assembly method applied for NHEJ integration, which made it challenging to construct and integrate metabolic pathways. In this study, a Golden Gate modular cloning system (YALIcloneNHEJ) was established to develop a robust DNA assembly platform in Y. lipolytica. By optimizing key factors, including the amounts of ligase and the reaction cycles, the assembly efficiency of 4, 7, and 10 fragments reached up to 90, 75, and 50%, respectively. This YALIcloneNHEJ system was subsequently applied for the overproduction of the sesquiterpene (-)-α-bisabolol by constructing a biosynthesis route and enhancing the flux in the mevalonate pathway. The resulting strain produced 4.4 g/L (-)-α-bisabolol, the highest titer reported in yeast to date. Our study expands the toolbox of metabolic engineering and is expected to enable a highly efficient production of various terpenoids.

Methods for Endoscopic Removal of Over-the-Scope Clip: A Systematic Review.

Aims: The over-the-scope clip (OTSC) has recently emerged as a new endoscopic device for treating gastrointestinal bleeding, perforations, fistulas, and leaks. A modified OTSC device (full-thickness resection device, FTRD) has been widely used for endoscopic full-thickness resection. However, there is less experience regarding the indications and methods for OTSC removal. We aimed to summarize the existing methods and indications for OTSC removal. Methods: We searched PubMed, Cochrane Library, and ClinicalTrials.gov to identify relevant publications on OTSC removal. The details of OTSC removal, including the methods, indications, success rates, adverse events, and failure causes, were extracted and summarized. A meta-analysis of pooled success rates was conducted using STATA 15.0. Results: Eighteen articles were included. The reported methods for OTSC removal included (1) grasping forceps, (2) the Nd : YAG laser, (3) argon plasma coagulation, (4) the remOVE system, (5) endoscopic mucosal resection/endoscopic submucosal dissection, and (6) ice-cold saline solution. Indications for OTSC removal were (1) poor healing, (2) OTSC misplacement, (3) repeat biopsy/therapy or further treatment, (4) adverse events after OTSC implantation, (5) removal after recovery, and (6) patient wishes. The pooled success rate of OTSC removal was 89% in patients treated with the remOVE system. Minor bleeding, superficial thermal damage, and superficial mucosal tears were common adverse events. Mucosal overgrowth was the main cause of OTSC removal failure. Conclusions: The remOVE system is the best investigated method, with sufficient efficacy and safety for OTSC removal. This is the first systematic review of OTSC removal and provides significant guidance for clinical practice.

Histone deacetylase-2 expression and activity in children with nephrotic syndrome with different glucocorticoid response.

BACKGROUND: Glucocorticosteroid (GC) is one of the most effective drugs available for the treatment of primary nephrotic syndrome (PNS) in children. However, some patients show little or no response to GC. The purpose of our research was to observe and describe the different levels of histone deacetylase-2 (HDAC2) expression in peripheral blood lymphocytes in children with PNS compared with various responses to the GC treatment, with the primary aim to assess the correlation between HDAC2 and GC resistance in PNS children. METHODS: Forty-eight patients with PNS suffering from their first attack prior to GC treatment were chosen as subjects. They were divided into two groups, those who had steroid-sensitive nephrotic syndrome (SSNS; n = 25) and those with steroid-resistant NS (SRNS; n = 23), according to their response to a 6-week course of oral prednisone. Twenty healthy children from the Physical Examination Center in the hospital served as the control group; Peripheral blood was collected at different time points prior to GC treatment and after regular therapy. RT-PCR, western blot, and enzyme-linked immunosorbent assay (ELISA) techniques were adopted to analyze HDAC2 mRNA, protein expression, and activity, respectively, in peripheral blood lymphocytes. The level of interleukin-8 (IL-8) in serum was measured by an ELISA. RESULTS: Prior to GC treatment, HDAC2 expression level and activity were lower in the SRNS group than in the SSNS and control group. A statistically significant difference in HDAC2 expression and activity were observed after GC treatment between these groups, with HDAC2 expression and activity lower in the SRNS group than in the SSNS and control groups. In the SSNS group, the expression and activity of HDAC2 were higher following GC treatment than prior to GC treatment. There was a clear difference in HDAC2 expression and activity of SRNS at the different time points. No statistically significant difference was found between the two groups. The pre-treatment and post-treatment serum IL-8 levels in the SRNS group were significantly higher than those in the SSNS group. HDAC2 from children with PNS before GC treatment and after regular therapy for 6 weeks was negatively correlated with serum IL-8 level. CONCLUSION: The GC effect was influenced by the HDAC2 expression and activity, leading to decreased serum IL-8 levels in children with PNS. HDAC2 seems to be one of the markers of GC resistance in children with PNS.

A genetic variant in vitamin B12 metabolic genes that reduces the risk of congenital heart disease in Han Chinese populations.

BACKGROUND: Genome-wide association studies on components of the one-carbon metabolic pathway revealed that human vitamin B12 levels could be significantly influenced by variations in the fucosyltransferase 2 (FUT2), cubilin (CUBN), and transcobalamin-I (TCN1) genes. An altered vitamin B12 level is an important factor that disturbs the homeostasis of the folate metabolism pathway, which in turn can potentially lead to the development of congenital heart disease (CHD). Therefore, we investigated the association between the variants of vitamin B12-related genes and CHD in Han Chinese populations. METHODS AND RESULTS: Six variants of the vitamin B12-related genes were selected for analysis in two independent case-control studies, with a total of 868 CHD patients and 931 controls. The variant rs11254363 of the CUBN gene was associated with a decreased risk of developing CHD in both the separate and combined case-control studies. Combined samples from the two cohorts had a significant decrease in CHD risk for the G allele (OR = 0.48, P = 1.7×10⁻5) and AG+GG genotypes (OR = 0.49, P = 4×10⁻5), compared with the wild-type A allele and AA genotype, respectively. CONCLUSIONS: Considering the G allele of variant rs11254363 of the CUBN gene was associated with an increased level of circulating vitamin B12. This result suggested that the carriers of the G allele would benefit from the protection offered by the high vitamin B12 concentration during critical heart development stages. This finding shed light on the unexpected role of CUBN in CHD development and highlighted the interplay of diet, genetics, and human birth defects.

Genetic variants reducing MTR gene expression increase the risk of congenital heart disease in Han Chinese populations.

AIMS: Elevated homocysteine levels are known to be a risk factor for congenital heart disease (CHD), but the mechanism underlying this effect is unknown. During early embryonic development, homocysteine removal is dictated exclusively by the MTR activity. To examine the role of MTR in CHD risk, we identified genetic variants in MTR and investigated the mechanisms that affect its expression levels and that increase the risk of CHD in Chinese populations. METHODS AND RESULTS: The association between regulatory variants of the MTR gene and CHD was examined in three independent case-control studies in a total of 2340 patients with CHD and 2270 controls. The functional consequences of these variants were demonstrated using dual-luciferase assays, real-time polymerase chain reaction (PCR), electrophoretic mobility shift assays, surface plasma resonance, chromatin immunoprecipitation, and bisulfite sequencing, as well as by a group of predicted microRNAs using a gene reporter system. Two regulatory variants of MTR, -186T>G and +905G>A, were associated with an increased risk of CHD in both the separate and combined case-control studies (-186GG P = 1.32 × 10(-9); +905AA P = 6.35 × 10(-14)). Compared with the major allele, the -186G allele exhibited significantly lower promoter activity, decreased hnRNA and mRNA levels, reduced transcription factor binding affinity, and a more highly methylated promoter. The +905A allele exhibited a statistically stronger binding affinity to functional microRNAs that down regulate MTR expression at the translational level. Both of the minor alleles were correlated with elevated plasma homocysteine concentrations, indicating a genetic component for hyperhomocysteinaemia. CONCLUSIONS: Regulatory variants of the MTR gene increase CHD risk by reducing MTR expression and inducing the homocysteine accumulation and elevation.

Physiological responses and evaluation of effects of BMI, smoking and drinking in high altitude acclimatization: a cohort study in Chinese Han young males.

High altitude acclimatization is a series of physiological responses taking places when subjects go to altitude. Many factors could influence these processes, such as altitude, ascending speed and individual characteristics. In this study, based on a repeated measurement design of three sequential measurements at baseline, acute phase and chronic phase, we evaluated the effect of BMI, smoking and drinking on a number of physiological responses in high altitude acclimatization by using mixed model and partial least square path model on a sample of 755 Han Chinese young males. We found that subjects with higher BMI responses were reluctant to hypoxia. The effect of smoking was not significant at acute phase. But at chronic phase, red blood cell volume increased less while respiratory function increased more for smoking subjects compared with nonsmokers. For drinking subjects, red blood cell volume increased less than nondrinkers at both acute and chronic phases, while blood pressures increased more than nondrinkers at acute phase and respiratory function, red blood cell volume and oxygen saturation increased more than nondrinkers at chronic phase. The heavy and long-term effect of smoking, drinking and other factors in high altitude acclimatization needed to be further studied.

Several polymorphisms of KCNQ1 gene are associated with plasma lipid levels in general Chinese populations.

BACKGROUND: Potassium voltage-gated channel, KQT-like subfamily, member 1 (KCNQ1) is thought to be an important candidate gene of diabetes. Several single nucleotide polymorphisms (SNPs) in a 40-kb linkage disequilibrium (LD) block in its intron 15 have been identified to be associated with diabetes in East Asian populations in recent genome-wide association studies. The aim of this study was to investigate whether KCNQ1 polymorphisms influence the levels of the metabolic phenotypes in general Chinese populations. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the associations of two SNPs (rs2237892 and rs2237895) in the aforementioned 40-kb LD block, a missense variant rs12720449 (P448R) in exon 10, and a synonymous variant rs1057128 (S546S) in exon 13 with metabolic phenotypes in a Uyghur population (n = 478) and replicated these associations in a Han population (n = 2,485). We found that rs2237892-T allele was significantly associated with decreased triglyceride levels (p(combined) = 0.001). The minor G allele of the rs12720449, with sharp difference of the allelic frequency between European and East Asian populations (0.2% versus 14%, respectively), was associated with a lower triglyceride levels than G allele in Uyghur subjects (p = 0.004), in Han subjects (p = 0.052), and in subjects of meta-analysis (p(combined) = 0.001). Moreover, the minor A allele of the rs1057128 was also associated with decreased triglyceride levels in meta-analysis (p(combined) = 0.010). CONCLUSIONS: To the best of our knowledge, this is the first report associating a missense mutation of KCNQ1, rs12720449, with triglyceride levels. Rs2237892, representing the 40-kb LD block, is also associated with triglyceride levels in Han population. Further studies are required to replicate these findings in other East Asian populations.