The m6A methyltransferase METTL3 affects cell proliferation and migration by regulating YAP expression in Hirschsprung disease.

BACKGROUND: METTL3, an mRNA m6A methyltransferase, has been implicated in various steps of mRNA metabolism, such as stabilization, splicing, nuclear transportation, translation, and degradation. However, whether METTL3 dysregulation is involved in Hirschsprung disease (HSCR) development remains unclear. In this study, we preliminarily elucidated the role of METTL3 in HSCR and sought to identify the associated molecular mechanism. METHODS: The gene expression levels of YAP and several methyltransferases, demethylases, and effectors were evaluated by RT-qPCR. Protein levels were evaluated by western blot and immunohistochemistry. Cell proliferation and migration were detected by CCK-8 and Transwell assays, respectively. The overall levels of m6A modification were determined by colorimetry. RESULTS: We found that m6A levels were reduced in the stenotic intestinal tissue of patients with HSCR. When METTL3 was knocked down in SH-SY5Y and HEK-293T cells, the proliferative and migratory abilities of the cells were inhibited, m6A modification levels were reduced, and YAP expression was increased. Importantly, YAP and METTL3 expression displayed a negative correlation in both cell lines as well as in HSCR tissue. CONCLUSIONS: Our results provide evidence for an interaction between METTL3 and YAP in HSCR, and further suggest that METTL3 is involved in the pathogenesis of HSCR by regulating neural crest cell proliferation and migration upstream of YAP.

Characterization of acetolactate decarboxylase of Streptococcus thermophilus and its stereoselectivity in decarboxylation of α-hydroxy-ß-ketoacids.

Acetolactate decarboxylase (ALDC) is a well-characterized catabolic enzyme catalyzes the decarboxylation of (±)-acetolactate to produce a single product, (R)-acetoin. It can also convert other racemic α-hydroxy-ß-ketoacids to corresponding α -hydroxyketones in R-configuration. In this work, we prepared ALDC of Streptococcus thermophilus (StALDC) and explored its stereoselectivity on different substrates. The enzyme displays no enantioselectivity on substrate (±)-acetolactate, but R-selectivity on product acetoin, which are identical with the data reported for various ALDCs. When compound (±)-2-propionyl-2-hydroxybutyrate is used as a substrate, however, the enzyme exhibits S-selectivity on both substrate and product, namely it can only decarboxylate (S)-2-propionyl-2-hydroxybutyrate to generate (S)-4-hydroxy-3-hexanone rather than its R-isomer, which is totally discriminate from the data published for the ALDC of Bacillus subtilis. As far as we know, this is the first time that substrate dependent enantioselectivity of ALDC is reported and the feature of StALDC is also discussed on the basis of homology modeling and molecular docking experiments.

Ambient climate determines the directional trend of community stability under warming and grazing.

Changes in ecological processes over time in ambient treatments are often larger than the responses to manipulative treatments in climate change experiments. However, the impacts of human-driven environmental changes on the stability of natural grasslands have been typically assessed by comparing differences between manipulative plots and reference plots. Little is known about whether or how ambient climate regulates the effects of manipulative treatments and their underlying mechanisms. We collected two datasets, one a 36-year long-term observational dataset from 1983 to 2018, and the other a 10-year manipulative asymmetric warming and grazing experiment using infrared heaters with moderate grazing from 2006 to 2015 in an alpine meadow on the Tibetan Plateau. The 36-year observational dataset shows that there was a nonlinear response of community stability to ambient temperature with a positive relationship between them due to an increase in ambient temperature in the first 25 years and then a decrease in ambient temperature thereafter. Warming and grazing decreased community stability with experiment duration through an increase in legume cover and a decrease in species asynchrony, which was due to the decreasing background temperature through time during the 10-year experiment period. Moreover, the temperature sensitivity of community stability was higher under the ambient treatment than under the manipulative treatments. Therefore, our results suggested that ambient climate may control the directional trend of community stability while manipulative treatments may determine the temperature sensitivity of the response of community stability to climate relative to the ambient treatment. Our study emphasizes the importance of the context dependency of the response of community stability to human-driven environmental changes.

Annual ecosystem respiration is resistant to changes in freeze-thaw periods in semi-arid permafrost.

Warming in cold regions alters freezing and thawing (F-T) of soil in winter, exposing soil organic carbon to decomposition. Carbon-rich permafrost is expected to release more CO2 to the atmosphere through ecosystem respiration (Re) under future climate scenarios. However, the mechanisms of the responses of freeze-thaw periods to climate change and their coupling with Re in situ are poorly understood. Here, using 2 years of continuous data, we test how changes in F-T events relate to annual Re under four warming levels and precipitation addition in a semi-arid grassland with discontinuous alpine permafrost. Warming shortened the entire F-T period because the frozen period shortened more than the extended freezing period. It decreased total Re during the F-T period mainly due to decrease in mean Re rate. However, warming did not alter annual Re because of reduced soil water content and the small contribution of total Re during the F-T period to annual Re. Although there were no effects of precipitation addition alone or interactions with warming on F-T events, precipitation addition increased total Re during the F-T period and the whole year. This decoupling between changes in soil freeze-thaw events and annual Re could result from their different driving factors. Our results suggest that annual Re could be mainly determined by soil water content rather than by change in freeze-thaw periods induced by warming in semi-arid alpine permafrost.

Microbial community responses reduce soil carbon loss in Tibetan alpine grasslands under short-term warming.

Changes in labile carbon (LC) pools and microbial communities are the primary factors controlling soil heterotrophic respiration (Rh ) in warming experiments. Warming is expected to initially increase Rh but studies show this increase may not be continuous or sustained. Specifically, LC and soil microbiome have been shown to contribute to the effect of extended warming on Rh . However, their relative contribution is unclear and this gap in knowledge causes considerable uncertainty in the prediction of carbon cycle feedbacks to climate change. In this study, we used a two-step incubation approach to reveal the relative contribution of LC limitation and soil microbial community responses in attenuating the effect that extended warming has on Rh . Soil samples from three Tibetan ecosystems-an alpine meadow (AM), alpine steppe (AS), and desert steppe (DS)-were exposed to a temperature gradient of 5-25°C. After an initial incubation period, soils were processed in one of two methods: (a) soils were sterilized then inoculated with parent soil microbes to assess the LC limitation effects, while controlling for microbial community responses; or (b) soil microbes from the incubations were used to inoculate sterilized parent soils to assess the microbial community effects, while controlling for LC limitation. We found both LC limitation and microbial community responses led to significant declines in Rh by 37% and 30%, respectively, but their relative contributions were ecosystem specific. LC limitation alone caused a greater Rh decrease for DS soils than AMs or ASs. Our study demonstrates that soil carbon loss due to Rh in Tibetan alpine soils-especially in copiotrophic soils-will be weakened by microbial community responses under short-term warming.

Fungal pathogens pose a potential threat to animal and plant health in desertified and pika-burrowed alpine meadows on the Tibetan Plateau.

Intact Tibetan meadows provide significant defense against soil-borne pathogen dispersal. However, dramatic meadow degradation has been observed due to climate change and pika damage, but their impacts on soil-borne pathogens are still unclear. With approximately 40% of the world's population living in Tibetan Plateau and its downstream watersheds, this lack of knowledge should be of great concern. Here, we used Illumina amplicon sequencing to characterize the changes in potential human, domestic animal, plant, and zoonotic bacterial and fungal pathogens in nondegraded, desertified, and pika-burrowed meadows. The relative abundance of bacterial domestic animal pathogens and zoonotic pathogens were significantly increased by desertification. Pika burrowing significantly increased the relative abundance of bacterial human pathogens and zoonotic pathogens. The species richness and relative abundance of fungal pathogens was significantly increased by desertification and pika burrowing. Accordingly, fungal plant and animal pathogens categorized by FUNGuid significantly increased in desertified and pika-burrowed meadows. Soil chemical and plant properties explained 38% and 64% of the bacterial and fungal pathogen community variance, respectively. Therefore, our study indicates for the first time that both alpine meadow desertification and pika burrowing could potentially increase infectious disease risks in the alpine ecosystem, especially for fungal diseases.

Glycyrrhizin, a High-Mobility Group Box 1 Inhibitor, Improves Lipid Metabolism and Suppresses Vascular Inflammation in Apolipoprotein E Knockout Mice.

BACKGROUND: High-mobility group box protein 1 (HMGB1) is known to have proinflammatory properties; however, the mechanisms by which HMGB1 influences immune responses during atherosclerosis (AS) development are not well understood. Thus, this study investigated the relationship between HMGB1 and vascular inflammation in Apoe-/- mice and whether glycyrrhizin (GLY), a small inhibitor of HMGB1, could have atheroprotective effects in AS. METHODS: Apoe-/- mice on a high-fat diet were treated with GLY (50 mg/kg) or vehicle by gavage once daily for 12 weeks, respectively. RESULTS: The GLY group exhibited significantly decreased serum lipid levels, atherosclerotic plaque deposition, and serum HMGB1 levels, as well as an increased Treg/Th17 ratio. The GLY group displayed increased interleukin-10 (IL-10) and IL-2 expression and decreased IL-17A and IL-6 expression. Furthermore, the GA treatment significantly reduced STAT3 phosphorylation in Th17 cells and increased STAT5 phosphorylation in Treg cells. CONCLUSIONS: Our findings indicate that the attenuation of atherosclerotic lesions in Apoe-/- mice by GLY might be associated with the amelioration of lipid metabolism abnormalities, inhibition of HMGB1 expression, and alterations in the Treg/Th17 ratio.

The Effects of High Mobility Group Box-1 Protein on Peripheral Treg/Th17 Balance in Patients with Atherosclerosis.

BACKGROUND: Atherosclerosis (AS) is defined as chronic inflammation of the vessel wall. The major objective of the this study was to explore the mechanism of Treg/Th17 imbalance and the role of high mobility group box-1 protein (HMGB1) on the balance in AS. METHODS: We detected the apoptotic ratios of Treg and Th17 cells in peripheral blood mononuclear cells (PBMCs) from subjects with AS and normal coronary arteries (NCA) by flow cytometry. The effects of recombinant HMGB1 (rHMGB1) on the proportion, apoptosis and differentiation of Treg and Th17 cells were analyzed using flow cytometry, qRT-PCR and ELISA. RESULTS: The frequencies of apoptotic Treg cells in the PBMCs from the subjects with AS were significantly higher than in those with NCA (p < 0.01). Stimulation of rHMGB1 obviously increased the level of Th17 cells and acid- related orphan receptor C (RORC) mRNA, and markedly decreased Treg cell frequency and the mRNA expression of factor forkhead family protein 3 (Foxp3) in the PBMCs. rHMGB1 played an obvious role in elevating Treg cell apoptosis ratio (p < 0.01). rHMGB1 treatment significantly decreased Treg cell ratio and IL-10 level, and increased Th17 cell ratio and IL-17A level induced from naïve CD4+ T cells. CONCLUSIONS: HMGB1 may modulate Treg/Th17 balance in patients with AS through inducing Treg cell apoptosis and promoting cell differentiation of Th17.

Plant Identity Exerts Stronger Effect than Fertilization on Soil Arbuscular Mycorrhizal Fungi in a Sown Pasture.

Arbuscular mycorrhizal (AM) fungi play key roles in plant nutrition and plant productivity. AM fungal responses to either plant identity or fertilization have been investigated. However, the interactive effects of different plant species and fertilizer types on these symbiotic fungi remain poorly understood. We evaluated the effects of the factorial combinations of plant identity (grasses Avena sativa and Elymus nutans and legume Vicia sativa) and fertilization (urea and sheep manure) on AM fungi following 2-year monocultures in a sown pasture field study. AM fungal extraradical hyphal density was significantly higher in E. nutans than that in A. sativa and V. sativa in the unfertilized control and was significantly increased by urea and manure in A. sativa and by manure only in E. nutans, but not by either fertilizers in V. sativa. AM fungal spore density was not significantly affected by plant identity or fertilization. Forty-eight operational taxonomic units (OTUs) of AM fungi were obtained through 454 pyrosequencing of 18S rDNA. The OTU richness and Shannon diversity index of AM fungi were significantly higher in E. nutans than those in V. sativa and/or A. sativa, but not significantly affected by any fertilizer in all of the three plant species. AM fungal community composition was significantly structured directly by plant identity only and indirectly by both urea addition and plant identity through soil total nitrogen content. Our findings highlight that plant identity has stronger influence than fertilization on belowground AM fungal community in this converted pastureland from an alpine meadow.

Enhanced photocatalytic performance of an Ag3PO4 photocatalyst via fullerene modification: first-principles study.

The coupling of carbon nanomaterials with semiconductor photocatalysts is a promising route to improve their photocatalytic performance. Herein, density functional theory was used to investigate the electronic structure, charge transfer, photocatalytic activity, and stability in a series of hybrid fullerene (C20, Li@C20, C26, Li@C26)/Ag3PO4(100) composites. When a Li atom is incorporated in fullerene, the adsorption energies significantly increase, although the change of interface distance is negligibly small due to the weak interface interaction. The charge transfer between constituents decreases with the C atom number of fullerene. Compared to pure Ag3PO4, the band gap of the composites is smaller, which enhances the visible-light absorption and photoinduced electron transfer. Most importantly, a type-II, staggered band alignment could be obtained in the C26-Ag3PO4(Li@C26-Ag3PO4) interface, leading to significantly reduced charge recombination and thus enhanced photocatalytic activity. These results reveal that fullerene modification would be an effective strategy to improve the photocatalytic performance of Ag3PO4 semiconductor photocatalysts.

Exploring effective sampling design for monitoring soil organic carbon in degraded Tibetan grasslands.

The effects of climate change and human activities on grassland degradation and soil carbon stocks have become a focus of both research and policy. However, lack of research on appropriate sampling design prevents accurate assessment of soil carbon stocks and stock changes at community and regional scales. Here, we conducted an intensive survey with 1196 sampling sites over an area of 190 km(2) of degraded alpine meadow. Compared to lightly degraded meadow, soil organic carbon (SOC) stocks in moderately, heavily and extremely degraded meadow were reduced by 11.0%, 13.5% and 17.9%, respectively. Our field survey sampling design was overly intensive to estimate SOC status with a tolerable uncertainty of 10%. Power analysis showed that the optimal sampling density to achieve the desired accuracy would be 2, 3, 5 and 7 sites per 10 km(2) for lightly, moderately, heavily and extremely degraded meadows, respectively. If a subsequent paired sampling design with the optimum sample size were performed, assuming stock change rates predicted by experimental and modeling results, we estimate that about 5-10 years would be necessary to detect expected trends in SOC in the top 20 cm soil layer. Our results highlight the utility of conducting preliminary surveys to estimate the appropriate sampling density and avoid wasting resources due to over-sampling, and to estimate the sampling interval required to detect an expected sequestration rate. Future studies will be needed to evaluate spatial and temporal patterns of SOC variability.

Case report: A case of epidermolysis bullosa complicated with pyloric atresia and a literature review.

Objective: This article aims to explore the diagnosis, molecular characteristics, treatment, and prognosis of epidermolysis bullosa with pyloric atresia (EB-PA). Methods: The clinical manifestations, diagnosis and treatment, and genetic characteristics of a patient with EB-PA admitted to our hospital were analysed. The disease subtypes, concomitant abnormalities, molecular characteristics, and prognosis of patients with EB-PA were summarized by searching the EB-PA-related literature since 2011. Results: We present a very low birth weight female infant with skin blisters and pyloric obstruction. Exome sequencing revealed heterozygous mutations in the ITGB4 gene: c.794dupC (p. S265fs*5) and c.2962G > A (p.A988T). This infant was diagnosed with EB-PA. Coverage of the wounds and Penicillin were used to prevent infection, but the patient eventually developed severe sepsis. A literature review was carried out including 49 cases of EB-PA; among these cases, 34 were preterm infants, weighing between 930 and 3,640 g. Of these EB-PA patients, 28 had accompanying malformations, including urinary system malformations and aplasia cutis congenita (ACC). Thirty-two patients identified the subtype of EB-PA, of whom 25 were diagnosed with junctional epidermolysis bullosa (JEB), 6 with epidermolysis bullosa simplex (EBS), and 1 with dystrophic epidermolysis bullosa (DEB). Genetic testing was conducted on 23 patients, of whom 15 carried Integrin Beta-4 (ITGB4) gene mutations and one JEB patient carried an Integrin Alpha-6 (ITGA6) gene mutation; 4 of the 5 EBS patients had Plectin (PLEC) gene mutations, and the other had an ITGB4 mutation. ITGB4 mutation cases involved 29 mutation sites, primarily concentrated in the region encoding the integrin beta subunit; PLEC mutation cases involved 7 mutation sites. Among all cases, 43 underwent pyloric atresia surgery, of whom 24 died postoperatively, and 6 without surgery therapy died within a short period. Conclusion: EB-PA is a rare genetic disorder characterized by increased skin fragility and PA involving mutations in the ITGB4, PLEC, or ITGA6 genes. EB-PA has a high incidence of complications and mortality, surgery and supportive therapy are currently the most common treatment options.

The effect of TEMPOL pretreatment on postoperative cognitive function, inflammatory response, and oxidative stress in aged rats under sevoflurane anesthesia.

INTRODUCTION: The heterocyclic compound 4-hydroxy-(2,2,6,6-Tetramethylpiperidin-1-yl)oxyl (TEMPOL) has a protective effect on neurological function in brain tissues damaged by ischemia and hypoxia. This study explored the effects of TEMPOL pretreatment on postoperative cognitive function in aged rats under sevoflurane anesthesia, focusing on inflammatory response and oxidative stress. METHODS: Sixty male rats were divided into normal control (C), sevoflurane anesthesia (S), TEMPOL pretreatment (T), and sevoflurane anesthesia + TEMPOL pretreatment (ST) groups (15 per group). Groups T and ST rats received continuous intraperitoneal TEMPOL (100 mg/kg) for 3 days, while groups C and S rats were injected with 0.9% saline. After pretreatment, groups S and ST received 3% sevoflurane anesthesia. RESULTS: Rats in group S exhibited a longer swimming distance, longer escape latency, lower frequency of platform crossing, and shorter dwell time in the targeted quadrant than those in groups C and T. Rats in group ST exhibited a shorter swimming distance, shorter escape latency, higher frequency of platform crossing, and longer dwell time in the targeted quadrant than those in group S. The expressions of interleukin-6, tumor necrosis factor-α, inducible nitric oxide synthase, and Ym1/2 messenger ribonucleic acid were higher in groups S and ST rats than in groups C and T rats and lower in group ST rats than in group S rat (p < .05). Superoxide dismutase (SOD), total antioxidant capacity (T-AOC), and glutathione peroxidase (GSH-Px) were lower, while malondialdehyde (MDA) was higher in groups S and ST rats than in groups C and T rats (p < .05). Group ST showed higher SOD, T-AOC, and GSH-Px, and lower MDA than group S (p < .05). CONCLUSIONS: TEMPOL pretreatment attenuated postoperative cognitive impairment induced by sevoflurane anesthesia in aged rats. This may be attributed to the downregulation of NR2B-CREB-BDNF pathway, reducing the inflammatory response and oxidative stress damage in hippocampal tissue.

Asymmetric warming reduces the strength of selection pressure of moderate grazing on reproductive phenology in alpine plants.

Both warming and grazing already affect the reproductive phenology of alpine plants. However, their effects have mostly been studied in isolation, and their interaction is still unclear. In this study, an asymmetric warming (average + 1.2 °C during daytime and + 1.7 °C during nighttime and + 1.5 °C during summer and + 2.0 °C during winter) with moderate grazing experiment was conducted for four years to determine their individual and interactive effects on the onsets and durations of reproductive phenophases for fifteen alpine plant species on the Qinghai-Tibetan Plateau. Individual warming and grazing simultaneously advanced the average start dates and ending dates of budding, flowering and fruiting by 5.3-6.2 days, and further resulted in smaller effects on their durations for most plant species. The interactions between warming and grazing on them varied with plant species and year, which advanced by average 12.1 days for all plant species. The effects of grazing on the temperature sensitivity of the start dates of reproductive phenophases (average by -8.5 days °C-1) were greater than that of warming alone (average by -3.4 days °C-1) and warming with grazing (average by -5.5 days °C-1) for most of the alpine plant species. There were significant effects of the previous phenological events on subsequent reproductive phenophases. Therefore, our results suggested that both warming and grazing advanced reproductive phenophases through altered soil temperature and soil moisture and carry-over effects of previous phenological events on subsequent phenological events. Warming reduced the temperature sensitivity of the start dates of reproductive phenophases to grazing, suggesting that it depressed strength of selection pressure of grazing on the onsets of reproductive phenology in alpine plants.

Warming delays but grazing advances leaf senescence of five plant species in an alpine meadow.

Leaf senescence is the final stage in the life cycle of leaves and is critical to plants' fitness as well as to ecosystem carbon and nutrient cycling. To date, most understanding about the responses of leaf senescence to environmental changes has derived from research in forests, but the topic has been relatively neglected, especially under grazing conditions, in natural grasslands. We conducted a 3-year manipulative asymmetric warming with moderate grazing experiment to explore the responses of leaf senescence of five main species in an alpine meadow on the Qinghai-Tibetan Plateau. We found that warming prolonged leaf longevity through earlier leaf-out and later leaf senescence, and grazing prolonged it through a greater advance in leaf-out than first leaf coloration for all plants. Warming did not affect leaf nitrogen (N) content or N resorption efficiency (NRE), but grazing increased N content in coloring leaves for P. anserine and P. nivea and decreased NRE for K. humilis, P. anserine under no-warming, and for P. nivea under warming. The interactive effects of warming and grazing on leaf phenology and leaf traits depended on species identity and year. There were positive relationships between leaf-out and leaf senescence mainly derived from grazing, and positive relationships between NRE from old leaves and leaf senescence for three out of five plant species. Therefore, our results indicated that earlier leaf-out could result in earlier leaf senescence only under grazing, but depending on plant species. Delayed leaf coloring increased NRE from old leaves for some plant species measured under warming and grazing. Our results suggested that alpine plants may develop strategies to adapt to warming and grazing to assimilate more carbon through prolonged leaf longevity rather than increased NRE through earlier leaf coloring in the alpine meadow.

Vegetation structural shift tells environmental changes on the Tibetan Plateau over 40 years.

Structural information of grassland changes on the Tibetan Plateau is essential for understanding alterations in critical ecosystem functioning and their underlying drivers that may reflect environmental changes. However, such information at the regional scale is still lacking due to methodological limitations. Beyond remote sensing indicators only recognizing vegetation productivity, we utilized multivariate data fusion and deep learning to characterize formation-based plant community structure in alpine grasslands at the regional scale of the Tibetan Plateau for the first time and compared it with the earlier version of Vegetation Map of China for historical changes. Over the past 40 years, we revealed that (1) the proportion of alpine meadows in alpine grasslands increased from 50% to 69%, well-reflecting the warming and wetting trend; (2) dominances of Kobresia pygmaea and Stipa purpurea formations in alpine meadows and steppes were strengthened to 76% and 92%, respectively; (3) the climate factor mainly drove the distribution of Stipa purpurea formation, but not the recent distribution of Kobresia pygmaea formation that was likely shaped by human activities. Therefore, the underlying mechanisms of grassland changes over the past 40 years were considered to be formation dependent. Overall, the first exploration for structural information of plant community changes in this study not only provides a new perspective to understand drivers of grassland changes and their spatial heterogeneity at the regional scale of the Tibetan Plateau, but also innovates large-scale vegetation study paradigm.

Effects of warming and grazing on soil N availability, species composition, and ANPP in an alpine meadow.

Uncertainty about the effects of warming and grazing on soil nitrogen (N) availability, species composition, and aboveground net primary production (ANPP) limits our ability to predict how global carbon sequestration will vary under future warming with grazing in alpine regions. Through a controlled asymmetrical warming (1.2/1.7 degrees C during daytime/nighttime) with a grazing experiment from 2006 to 2010 in an alpine meadow, we found that warming alone and moderate grazing did not significantly affect soil net N mineralization. Although plant species richness significantly decreased by 10% due to warming after 2008, we caution that this may be due to the transient occurrence or disappearance of some rare plant species in all treatments. Warming significantly increased graminoid cover, except in 2009, and legume cover after 2008, but reduced non-legume forb cover in the community. Grazing significantly decreased cover of graminoids and legumes before 2009 but increased forb cover in 2010. Warming significantly increased ANPP regardless of grazing, whereas grazing reduced the response of ANPP to warming. N addition did not affect ANPP in both warming and grazing treatments. Our findings suggest that soil N availability does not determine ANPP under simulated warming and that heavy grazing rather than warming causes degradation of the alpine meadows.

Methanotrophic community structure and activity under warming and grazing of alpine meadow on the Tibetan Plateau.

Knowledge about methanotrophs and their activities is important to understand the microbial mediation of the greenhouse gas CH(4) under climate change and human activities in terrestrial ecosystems. The effects of simulated warming and sheep grazing on methanotrophic abundance, community composition, and activity were studied in an alpine meadow soil on the Tibetan Plateau. There was high abundance of methanotrophs (1.2-3.4 × 10(8) pmoA gene copies per gram of dry weight soil) assessed by real-time PCR, and warming significantly increased the abundance regardless of grazing. A total of 64 methanotrophic operational taxonomic units (OTUs) were obtained from 1,439 clone sequences, of these OTUs; 63 OTUs (98.4%) belonged to type I methanotrophs, and only one OTU was Methylocystis of type II methanotrophs. The methanotroph community composition and diversity were not apparently affected by the treatments. Warming and grazing significantly enhanced the potential CH(4) oxidation activity. There were significantly negative correlations between methanotrophic abundance and soil moisture and between methanotrophic abundance and NH(4)-N content. The study suggests that type I methanotrophs, as the dominance, may play a key role in CH(4) oxidation, and the alpine meadow has great potential to consume more CH(4) under future warmer and grazing conditions on the Tibetan Plateau.

[Discussion on methods for locating Xuehai (SP10) acupoint].

The position of Xuehai (SP10) is clear, but its locating method is vague, resulting in the disunity of clinical application and even possibly affecting the curative effect. Also, when learning the meridians and acupoints, the beginners are often confused by this issue possibly due to: ① when the bone-length proportional measurement combined with anatomic symbol (combination method) was adopted, it is not clear that the patient should take a posture of knee extension or knee flexion; ② when the combination method used, it is difficult to find the highest point of muscle eminence in the case of patient with thin vastus medialis muscle and fuzzy body surface projection; ③the simple method for locating SP10 is widely used at present, can it replace the combination method to locate this acupoint accurately？Guided by these questions, we, in the present paper, reviewed the rela-ted textbooks, works and other literature to explore the standard position of SP10, and the standard and simple methods for locating this acupoint. Comprehending various opinions, we hold that SP10 should be positioned under the extended knee posture, then, the acupoint's horizontal ordinate "2 cun superior to the medial end of the base of the patella" is determined by using bone-length proportional measurement to measure 2 cun from the bottom to the tip of the patella. Then, the body surface anatomic symbol method is used, when, the patient is asked to stretch the leg and contract the vastus medialis muscle, the highest spot of muscular eminence is the SP10. If the patient's muscular protuberance is not obvious, the middle line between the medial and lateral margins of the vastus medialis muscle is used as the vertical ordinate, and its intersection with the abscissa is SP10. The simple method is easy in operation and has a reference value, but may frequently produce errors, hence, it is not a substitution for the combination method.

Association of rs2435357 and rs2506030 polymorphisms in RET with susceptibility to hirschsprung disease: A systematic review and meta-analysis.

Background: There are numerous published studies on the association between RET polymorphisms and susceptibility to Hirschsprung disease (HSCR). However, some of the results are inconsistent and the studies were conducted with small sample sizes. Therefore, we performed a meta-analysis to clarify the relationship. Methods: Relevant data were retrieved from PubMed, Web of Science, Cochrane Library, EMBASE, CNKI, and Google Scholar according to PRISMA guidelines. Odds ratios (OR) were calculated to assess susceptibility to HSCR. Meanwhile, heterogeneity and publication bias were also calculated by R software package (version 4.2.1). The protocol was published in PROSPERO (CRD42022348940). Results: A total of 12 studies were included in the meta-analysis and comprised 12 studies on the RET polymorphism rs2435357 (1,939 subjects and 3,613 controls) and 7 studies on the RET polymorphism rs2506030 (1,849 patients with HSCR and 3,054 controls). The analysis revealed that rs2435357 [A vs. G: odds ratio (OR) = 3.842, 95% confidence interval (CI) 2.829-5.220; AA vs. GG: OR = 2.597, 95% CI 1.499-4.501; AA + AG vs. GG: OR = 6.789, 95% CI 3.0711-14.9973; AA vs. AG + GG: OR = 8.156, 95%CI 5.429-12.253] and rs2506030 (A vs. G: OR = 0.519, 95% CI 0.469-0.573; AA vs. GG: OR = 0.543, 95% CI 0.474-0.623; AA + AG vs. GG: OR = 0.410, 95% CI 0.360-0.468; AA vs. AG + GG: OR = 0.361, 95%CI 0.292-0.447) were significantly associated with susceptibility to HSCR. Conclusions: The polymorphisms rs2435357 and rs2506030 in the RET may be related to susceptibility to HSCR, of which rs2435357 (T > C) is the causal locus and rs2506030 (A > G) is the protective locus. Systematic Review Registration: https://www.crd.york.ac.uk/prospero/, identifier:CRD42022348940.