碘造影剂致急性血小板减少1例

分析1例住院患者全腹部增强CT检查时,使用碘造影剂40 min后出现过敏反应并伴血小板一过性急剧下降的原因,考虑可能是造影剂碘克沙醇过敏所致。本案例为临床提供预警作用,临床上应对此类情况患者进行积极预防和治疗。     

SRS-FISH: A high-throughput platform linking microbiome metabolism to identity at the single-cell level

One of the biggest challenges in microbiome research in environmental and medical samples is to better understand functional properties of microbial community members at a single-cell level. Single-cell isotope probing has become a key tool for this purpose, but the current detection methods for determination of isotope incorporation into single cells do not allow high-throughput analyses. Here, we report on the development of an imaging-based approach termed stimulated Raman scattering–two-photon fluorescence in situ hybridization (SRS-FISH) for high-throughput metabolism and identity analyses of microbial communities with single-cell resolution. SRS-FISH offers an imaging speed of 10 to 100 ms per cell, which is two to three orders of magnitude faster than achievable by state-of-the-art methods. Using this technique, we delineated metabolic responses of 30,000 individual cells to various mucosal sugars in the human gut microbiome via incorporation of deuterium from heavy water as an activity marker. Application of SRS-FISH to investigate the utilization of host-derived nutrients by two major human gut microbiome taxa revealed that response to mucosal sugars tends to be dominated by Bacteroidales, with an unexpected finding that Clostridia can outperform Bacteroidales at foraging fucose. With high sensitivity and speed, SRS-FISH will enable researchers to probe the fine-scale temporal, spatial, and individual activity patterns of microbial cells in complex communities with unprecedented detail.

With the rapid advances in both genotyping and phenotyping of single cells, bridging genotype and phenotype at the single-cell level is becoming a new frontier of science (1). Methods have been developed to shed light on the genotype–metabolism relationship of individual cells in a complex environment (2, 3), which is especially relevant for an in-depth understanding of complex microbial communities in the environment and host-associated microbiomes. For functional analyses of microbial communities, single-cell isotope probing is often performed in combination with nanoscale secondary ion mass spectrometry (NanoSIMS) (4–7), microautoradiography (MAR) (8, 9), or spontaneous Raman microspectroscopy (10–12) to visualize and quantify the incorporation of isotopes from labeled substrates. These methods can be combined with fluorescence in situ hybridization (FISH) using ribosomal ribonucleic acid (rRNA)-targeted probes (13), enabling a direct link between metabolism and identity of the organisms. In addition, Raman-activated cell sorting has been recently developed using either optical tweezers or cell ejection for downstream sequencing of the sorted cells (14–16). While these approaches have expanded the possibilities for functional analyses of microbiome members (17), all of the aforementioned methods suffer from extremely limited throughput. Consequently, only relatively few samples and cells per sample are typically analyzed in single-cell stable isotope probing studies, hampering a comprehensive understanding of the function of microbes in their natural environment.To overcome the limited throughput of Raman spectroscopy, coherent Raman scattering microscopy based on coherent anti-Stokes Raman scattering (CARS) or stimulated Raman scattering (SRS) has been developed (18, 19). Compared with CARS, the SRS signal is free of the electronic resonance response (20) and is linear to molecular concentration, thus permitting quantitative mapping of biomolecules (21, 22). Both CARS and SRS microscopy have successfully been applied for studying single-cell metabolism in eukaryotes (23–26). In a label-free manner, SRS imaging has led to the discovery of an aberrant cholesteryl ester storage in aggressive cancers (27, 28), lipid-rich protrusions in cancer cells under starvation (29), and fatty acid unsaturation in ovarian cancer stem cells (30) and more recently, in melanoma (31, 32). CARS and SRS have also been harnessed to explore lipid metabolism in live Caenorhabditis elegans (33–36). Combined with stable isotope probing, SRS microscopy has allowed the tracing of glucose metabolism in eukaryotic cells (37, 38) and the visualization of metabolic dynamics in living animals (25). Recently, SRS was successfully applied to infer antibiotic resistance patterns of bacterial pure cultures and heavy water (D2O) metabolism (39). Yet, SRS microscopy has not been adapted for studying functional properties of members of microbiomes as SRS itself lacks the capability of identifying cells in a complex community.Here, we present an integrative platform that exploits the advantages of SRS for single-cell stable isotope probing together with two-photon FISH for the identification of cells in a high-throughput manner. To deal with the challenges in detecting low concentrations of metabolites inside small cells with diameters around 1 µm, we have developed a protocol that maximizes the isotope label content in cells and exploits the intense SRS signal from the Raman band used for isotope detection.Conventionally, FISH is performed separately by one-photon excited fluorescence microscopy (40). To enhance efficiency, we developed a system that implements highly sensitive SRS metabolic imaging with two-photon FISH using the same laser source. These efforts collectively led to a high-throughput platform that enables correlative imaging of cell identity and metabolism at a speed of 10 to 100 ms per cell. In comparison, it takes about 20 s to record a Raman spectrum from a single cell in a conventional spontaneous Raman FISH experiment (41, 42).Our technology enabled high-throughput analysis of single-cell metabolism in the human gut microbiome. In the human body, microbes have been shown to modulate the host’s health (43, 44). Analytical techniques looking into their activities and specific physiologies (i.e., phenotype) as a result of both genotype and the environment provide key information on how microbes function, interact with, and shape their host. As a proof of principle, we used stimulated Raman scattering–two-photon fluorescence in situ hybridization (SRS-FISH) to track the incorporation of deuterium (D) from D2O into a mixture of two distinct gut microbiota taxa. Incorporation of D from D2O into newly synthesized cellular components of active cells, such as lipids and proteins, occurs analogously to incorporation of hydrogen from water during the reductive steps of biosynthesis of various cellular molecules (10, 45, 46). Importantly, D incorporation from D2O has been shown to be reliable to track metabolic activity of individual cells within complex microbial communities in response to the addition of external substrates (10, 17, 47). When microbial communities are incubated in the presence of D2O under nutrient-limiting conditions, individual cells display only minimal activity and only minor D incorporation (11, 17, 47). In contrary, when cells are stimulated by the addition of an external nutrient, cells that can metabolize this compound become active and incorporate D into macromolecules, which lead to the presence of C-D bonds into the cell’s biomass. Consequently, D incorporation from D2O can be combined with techniques able to detect C-D signals, such as Raman-based approaches, and to track metabolic activity at the single-cell level in response to a variety of compounds. Here, we show that SRS-FISH enables fast and sensitive determination of the D content of individual cells while simultaneously unveiling their phylogenetic identity. We applied this technique to complex microbial communities by tracking in situ the metabolic responses of two major phylogenetic groups of microbes in the human gut (Bacteroidales and Clostridia spp.) and of a particular species within each group to supplemented host-derived nutrients. Our study revealed that 1) Clostridia spp. can actually outperform Bacteroidales spp. at foraging on the mucosal sugar fucose and shows 2) a significant interindividual variability of responses of these major microbiome taxa toward mucosal sugars. Together, our results demonstrate the capability of SRS-FISH to unveil the metabolism of particular microbes in complex communities at a throughput that is two to three orders of magnitude higher than other metabolism identity bridging tools, therefore providing a valuable multimodal platform to the field of single-cell analysis.     

The homeostatic malfunction of a novel feedback pathway formed by lncRNA021545, miR-330-3p and epiregulin contributes in hepatocarcinoma progression via mediating epithelial-mesenchymal transition

A better understanding of tumor metastasis is urgently required for the treatment and prognosis of hepatocarcinoma patients. Current work contributes a novel ceRNA feedback regulation pathway composed of epiregulin (EREG), microRNA-330-3p (miR-330-3p) and long non-coding RNA 021545 (lncRNA021545) in regulating hepatocarcinoma malignancy via epithelial-mesenchymal transition (EMT) process. Closely correlated, the deficiencies of EREG and lncRNA021545 and the overexpression of miR-330-3p were involved in the clinical progression of hepatocarcinoma. In vitro results showed that 1) lncRNA021545 downregulation promoted, 2) miR-330-3p dysexpression positively correlated, and 3) EREG dysexpression reversely correlated with the migratory and invasive properties of hepatocarcinoma HCCLM3 and Huh7 cell lines. By directly binding to EREG and lncRNA021545, miR-330-3p expression change reversely correlated with their expressions in HCCLM3 and Huh7 cells, which was also confirmed in primary tumors from HCCLM3-xenograft mice in responding to miR-330-3p change. LncRNA021545 and EREG positively regulated each other, and lncRNA021545 negatively regulated miR-330-3p, while, EREG dysregulation unchanged miR-330-3p expression in hepatocarcinoma cells. Furthermore, systemic in vitro cellular characterizations showed that the malfunctions of the three molecules mediated the invasiveness of hepatocarcinoma cells via EMT process through affecting the expressions of E-cadherin, N-cadherin, vimentin, snail and slug, which was further confirmed by in vivo miR-330-3p promotion on the tumorigenicity and metastasis of HCCLM3 bearing nude mice and by in vitro miR-330-3p promotion on the migration and invasion of hepatocarcinoma cells to be antagonized by EREG overexpression through acting on EMT process. Our work indicates, that by forming a circuit signaling feedback pathway, the homeostatic expressions of lncRNA021545, miR-330-3p and EREG are important in liver health. Its collapse resulted from the downregulations of lncRNA021545 and EREG together with miR-330-3p overexpression promote hepatocarcinoma progression by enhancing the invasiveness of tumor cells through EMT activation. These discoveries suggest that miR-330-3p/lncRNA021545/EREG axis plays a critical role in hepatocarcinoma progression and as a candidate for its treatment.     

Trends in Physical Fitness and Nutritional Status among School-Aged Children and Adolescents during the COVID-19 Pandemic in Shaanxi,China—A Cross-Sectional Study

Background: This study aimed to explore the characteristics of trends in physical fitness and nutritional status among school-aged students during the COVID-19 pandemic, which could help to develop targeted guidelines and policies for students (adolescents) to promote health during lockdowns resulting from public health emergencies. Methods: The physical fitness and nutritional status were measured from 2019 to 2021; the correlation between years and grade were analyzed. Results: a declining trend was found in aerobic fitness, strength fitness, speed fitness, and BMI during this lockdown. Conclusion: The changes in dietary behavior and the decrease in outdoor physical activities may be the reason for the decline. Furthermore, differences in subjects’ growth and limited space at home must be considered in the formulation of exercise and nutritional plans. According to the results of our study, exercise for aerobic fitness should receive more attention when students are enduring a pandemic lockdown. In addition, saying “no” to high-calorie foods in the form of snacking and ultra-processed food was the key to improving subjects’ nutritional status.     

Study on the Initiation of Interface Crack in Rock Joints

The interfacial fracture of rock joints is an important although easily ignored issue in jointed rock engineering. To conduct this study, an interface crack model of rock joints was proposed. By analyzing the ratio of stress intensity factor to fracture toughness, the fracture mode of the interface crack was studied. Based on the Mohr-Coulomb criterion, an interface fracture criterion considering T-stress was established. To verify the proposed fracture criterion, laboratory and numerical tests were conducted. Finally, the effect of relative critical size α, internal friction angle φ and cohesion c on the initiation of an interface crack was comprehensively discussed. It is concluded that the proposed fracture criterion can predit the initiation of the interface cracks properly. With an increase in cohesion c, mode II fracture toughness K_IIC also clearly increases. When the absolute value of K_I is small, the effect of α is much larger than that of φ. In addition, with an increase in the absolute value of the mode I stress intensity factor, the φ of the joint plays a more important role in the initiation of the interface crack.     

亚砷酸对人肝癌细胞生长增殖及分化的影响

目的 探讨岖砷酸对人肝癌细胞BEL-7402增殖、分化的作用。方法 通过体外细胞培养,用四氮唑盐(MTT)比色法观察细胞生长曲线的变化,台盼蓝拒染法观察细胞群体倍增时间(TD)的改变,并分别用自动化学发光法、生化法检测亚砷酸对细胞甲胎蛋白(AFP)合成及谷氨酰转肽酶(GGT)、乳酸脱氢酶(LDH)活性的影响,用苏木精-伊红(HE)染色法及透射电镜观察细胞的分化形态特点。结果 亚砷酸(1．0～8．0μmol／L)能抑制BEL-7402细胞生长增殖,呈剂量、时间依赖性。亚砷酸(8．0μmoL／L)对BEL-7402细胞AFP合成及GGT、LDH活性有显著的抑制作用,HE染色及透射电镜观察可见部分细胞呈现分化的形态改变。结论 亚砷酸体外能够抑制人肝癌细胞生长增殖,并具有一定的诱导分化作用。     

Hsa_circ_0007142 contributes to cisplatin resistance in esophageal squamous cell carcinoma via miR‐494‐3p/LASP1 axis

BackgroundChemoresistance is one of the major obstacles for tumor treatment. Circular RNAs (circRNAs) have been confirmed to play vital roles in chemoresistance of cancer, including esophageal squamous cell carcinoma (ESCC). We investigated the roles and mechanisms of circ_0007142 in cisplatin (DDP) resistance of ESCC.MethodsQuantitative real‐time polymerase chain reaction (qRT‐PCR) was conducted to determine the levels of circ_0007142, DOCK1 mRNA, microRNA‐494‐3p (miR‐494‐3p) and LIM And SH3 Protein 1 (LASP1) mRNA. RNase R assay was conducted to analyze the characteristic of circ_0007142. Cell Counting Kit‐8 (CCK‐8) assay was performed to evaluate IC50 of DDP. Flow cytometry analysis, 5‐ethynyl‐2’‐deoxyuridine (EdU) assay and transwell assay were carried out to examine cell apoptosis, proliferation and invasion, respectively. Dual‐luciferase reporter assay was employed to verify the association between miR‐494‐3p and circ_0007142 or LASP1. Murine xenograft assay was conducted to investigate the role of circ_0007142 in DDP resistant in vivo. The protein level of LASP1 in tumors was measured by Immunohistochemistry (IHC) analysis.ResultsCirc_0007142 was upregulated in DDP‐resistant ESCC tissues and cells. Circ_0007142 knockdown improved DDP sensitivity, induced cell apoptosis and hampered cell proliferation and invasion in DDP‐resistant ESCC cells. Circ_0007142 functioned as the sponge for miR‐494‐3p and miR‐494‐3p inhibition reversed the impacts of circ_0007142 knockdown on DDP resistance, cell apoptosis, proliferation, and invasion. LASP1 was a target of miR‐494‐3p, and the effects on DDP resistance, cell apoptosis, growth, and invasion mediated by LASP1 downregulation were rescued by miR‐494‐3p inhibition. Moreover, circ_0007142 knockdown enhanced DDP sensitivity in vivo.ConclusionCirc_0007142 improved DDP resistance of ESCC by upregulating LASP1 via sponging miR‐494‐3p.     

重和林M30在新诊2型糖尿病中的应用

目的：观察重和林M 30（精蛋白重组人胰岛素30／70）在新诊2型糖尿病人中的临床作用。方法选取80例需胰岛素治疗的2型糖尿病人,随机分为重和林M 30组（试验组）与门冬胰岛素30组（对照组）,连续治疗8周,比较两组治疗前后空腹血糖（FBG）、餐后2 h血糖（PBG）、体重指数（BMI）、糖化血红蛋白（HbA1c）、空腹C肽（Fasting C peptide ,FCP）变化,评估低血糖、严重低血糖发生率及胰岛素用量和性价比。结果两组患者FBG、PBG、HbA1c、FCP较治疗前均有明显下降（P＜0.05）,两组患者治疗后的FBG、BMI、HbA1c、FCP比较无明显差异（P＞0.05）;两组患者低血糖发生率及胰岛素用量无明显差异（ P＞0.05 ）,两组治疗后均未见严重的低血糖反应,试验组性价比较高。结论重和林M 30与门冬胰岛素30治疗新诊2型糖尿病整体降糖疗效无明显差异,重和林M 30性价比较高,在基层医院具有广泛应用优势。