Causal relationships between circulating inflammatory cytokines and diabetic neuropathy: A Mendelian Randomization study.

BACKGROUND: Emerging evidence suggests systemic inflammation as a critical mechanism underlying diabetic neuropathy. This study aimed to investigate the causal relationship between 41 circulating inflammatory cytokines and diabetic neuropathy. METHODS: Summary statistics from previous Genome-Wide Association studies (GWAS) included pooled data on 41 inflammatory cytokines and diabetic neuropathy. A two-sample Mendelian Randomization (MR) design was employed, and the robustness of the results was confirmed through comprehensive sensitivity analyses. RESULTS: Our study reveals that the linkage between increased levels of IFN_G (OR = 1.31, 95 %CI: 1.06-1.63; P = 0.014), IP_10 (OR = 1.18, 95 %CI: 1.01-1.36; P = 0.031) and an elevated risk of diabetic neuropathy. Conversely, higher levels of IL_9 (OR = 0.86, 95 %CI: 0.75-1.00; P = 0.048) and SCF (OR = 0.83, 95 %CI: 0.73-0.94; P = 0.003) are genetically determined to protect against diabetic neuropathy. Furthermore, the sensitivity analysis affirmed the results' dependability, revealing no heterogeneity or pleiotropy. CONCLUSION: Our MR research identified four upstream inflammatory cytokines implicated in diabetic neuropathy. Overall, these findings suggest the potential for innovative therapeutic strategies. Further large-scale cohort studies are required for validation.

Rapid and sensitive liquid chromatography with tandem mass spectrometry method for the simultaneous quantification of yonkenafil and its major metabolites in rat plasma.

Yonkenafil is a promising drug for treatment of male erectile dysfunction. Previous studies showed that the piperazine-N,N'-deethylation metabolite, piperazine-N-deethylation metabolite, and piperazine-N-deethylation-N,N'-deethylation metabolite were the major metabolites of yonkenafil after extensive metabolism. We developed a sensitive and selective method for the simultaneous quantification of yonkenafil and its major metabolites using high-throughput liquid chromatography with tandem mass spectrometry. Analytes and internal standard were extracted from a small quantity of plasma (50 µL) using liquid-liquid extraction with diethyl ether/dichloromethane (60:40, v/v), and the baseline separation was achieved on Zorbax SB-C18 column using ammonia/water/methanol (0.2:20:80, v/v/v) as the mobile phase. The assay was performed with an electrospray positive ionization mass spectrometry through the multiple-reaction monitoring mode within 2 min. Calibration curve of the method was linear within the range of 1.00-1000 ng/mL for all the analytes with the intra- and interday precisions of 4.0-5.2 and 4.0-5.3% for yonkenafil, 3.1-4.9 and 3.1-5.2% for the piperazine-N,N'-deethylation metabolite, 4.8-6.8 and 4.8-7.3% for the piperazine-N-deethylation metabolite, and 2.9-6.1 and 5.4-6.3% for the piperazine-N-deethylation-N,N'-deethylation metabolite, respectively. The recoveries were above 90% with low matrix effects. The validated assay was successfully applied to support a preclinical pharmacokinetic study in six rats using a single oral dose of yonkenafil (8 mg/kg).

Determination of thymopentin in beagle dog blood by liquid chromatography with tandem mass spectrometry and its application to a preclinical pharmacokinetic study.

The pentapeptide thymopentin (Arg-Lys-Asp-Val-Tyr, RKDVY) corresponds to amino acids 32-36 of the 49 amino acid immunomodulatory polypeptide, thymopoietin, whose biological activity is partially reproduced. Thymopentin is widely used in the clinic and represents a promising target for drug design but bioanalytical and pharmacokinetic data are limited due to its enzymatic instability. This paper reports a rapid and sensitive method based on liquid chromatography with tandem mass spectrometry for the determination of thymopentin in beagle dog blood. To inactivate peptidases and stabilize thymopentin, acetonitrile was added to blood samples immediately after collection followed by addition of stable isotope-labeled thymopentin as internal standard and washing with dichloromethane. Chromatography was carried out on an Ascentis Express Peptide ES-C18 column using gradient elution with methanol and aqueous 0.1% formic acid at a flow rate of 0.6 mL/min. Positive electrospray ionization mass spectrometry with selected reaction monitoring achieved linearity in the range of 1.5-800 ng/mL with good accuracy/precision and minimal matrix effects. The method was successfully applied to a pharmacokinetic study in beagle dogs after intravenous administration of 0.2 mg/kg thymopentin.

Causal relationships between gut microbiota, gut metabolites, and diabetic neuropathy: A mendelian randomization study.

BACKGROUND: Previous studies have shown a strong correlation between gut microbiota and diabetes and its associated complications. We aimed to evaluate the causal relationships between the gut microbiota, gut metabolites, and diabetic neuropathy. METHODS: Summary statistics of 211 gut microbiota and 12 gut-related metabolites (ß-hydroxybutyric acid, betaine, trimethylamine-N-oxide, carnitine, choline, glutamate, kynurenine, phenylalanine, propionic acid, serotonin, tryptophan, and tyrosine) were obtained from previous genome-wide association studies (GWAS). A two-sample Mendelian randomization (MR) design was used to estimate the effects of gut microbiota and gut metabolites on the risk of diabetic neuropathy based on FinnGen GWAS. RESULTS: Higher levels of Acidaminococcaceae (OR = 0.62; 95%CI = 0.46 to 0.84; P = 0.002), Peptococcaceae (OR = 0.70; 95%CI = 0.54 to 0.90; P = 0.006), and Eubacterium coprostanoligenes group (OR = 0.68; 95%CI = 0.50 to 0.93; P = 0.016) are genetically determined to provide protection against diabetic neuropathy. Conversely, the presence of Alistipes (OR = 1.65; 95%CI = 1.18 to 2.31; P = 0.003), ChristensenellaceaeR7 group (OR = 1.52; 95%CI = 1.03 to 2.23; P = 0.033), Eggerthella (OR = 1.28; 95%CI = 1.05 to 1.55; P = 0.014), RuminococcaceaeUCG013 (OR = 1.35; 95%CI = 1.01 to 1.82; P = 0.046), and Firmicutes (OR = 1.42; 95%CI = 1.05 to 1.93; P = 0.023) increases the risk of diabetic neuropathy. Moreover, a correlation has been identified between diabetic neuropathy and two gut metabolites: betaine (OR = 0.95; 95%CI = 0.90 to 1.00; P = 0.033) and tyrosine (OR = 1.03; 95%CI = 1.01 to 1.06; P = 0.019). Sensitivity analysis indicated robust results with no sign of heterogeneity or pleiotropy. CONCLUSION: The present study elucidated the impact of specific gut microbiota and gut metabolites on the susceptibility to diabetic neuropathy. Interventions targeting the improvement of the gut microbiota diversity and composition hold considerable promise as a potential strategy.

Observation and research of deep underground multi-physical fields-Huainan -848 m deep experiment.

Compared with the surface, the deep environment has the advantages of allowing "super-quiet and ultra-clean"-geophysical field observation with low vibration noise and little electromagnetic interference, which are conducive to therealization of long-term and high-precision observation of multi-physical fields, thus enabling the solution of a series of geoscience problems. In the Panyidong Coal Mine, where there are extensive underground tunnels at the depth of 848 m belowsea level, we carried out the first deep-underground geophysical observations, including radioactivity, gravity, magnetic, magne-totelluric, background vibration and six-component seismic observations. We concluded from these measurements that (1) the background of deep subsurface gravity noise in the long-period frequency band less than 2 Hz is nearly two orders ofmagnitude weaker than that in the surface observation environment; (2) the underground electric field is obviously weaker thanthe surface electric field, and the relatively high frequency of the underground field, greater than 1 Hz, is more than two orders of magnitude weaker than that of the surface electric field; the east-west magnetic field underground is approximately the same asthat at the surface; the relatively high-frequency north-south magnetic field underground, below 10 Hz, is at least one order ofmagnitude lower than that at the surface, showing that the underground has a clean electromagnetic environment; (3) in additionto the high-frequency and single-frequency noises introduced by underground human activities, the deep underground spacehas a sig-nificantly lower background vibration noise than the surface, which is very beneficial to the detection of weakearthquake and gravity signals; and (4) the underground roadway support system built with ferromagnetic material interferesthe geomagnetic field. We also found that for deep observation in the "ultra-quiet and ultra-clean" environment, the existinggeophysical equipment and observation technology have problems of poor adaptability and insufficient precision as well asdata cleaning problems, such as the effective separation of the signal and noise of deep observation data. It is also urgent tointerpret and comprehensively utilize these high-precision multi-physics observation data. Electronic Supplementary Material: Supplementary material is available in the online version of this article at 10.1007/s11430-022-9998-2.

Association between thyroid hormones and skeletal muscle and bone in euthyroid type 2 diabetes patients.

Background: The impact of thyroid hormones within their normal ranges on skeletal muscle and bone in patients with type 2 diabetes mellitus (T2DM) remains unknown. The purpose of this study was to investigate the relationships of thyroid hormones with muscle and bone in euthyroid patients with T2DM. Methods: This cross-sectional study included 344 euthyroid T2DM patients. Muscle mass and bone mineral density were measured by dual-energy X-ray absorptiometry. The levels of thyroid-stimulating hormone (TSH), free triiodothyronine (FT3), and free thyroxin (FT4) were measured by electrochemiluminescence immunoassay. Results: The results revealed that FT3 was positively correlated with body mass index (BMI) in male patients after age correction. In men, FT4 was negatively correlated with body weight, BMI, total muscle mass, appendicular skeletal muscle mass (ASM), and ASM index (ASMI), while FT3/FT4 was positively correlated with body weight, BMI, total muscle mass, ASM, and ASMI after age correction. In women, FT4 was negatively correlated with ASM and ASMI, while FT3/FT4 was positively correlated with ASM and ASMI after age correction. FT3/FT4 was significantly lower in men with low muscle mass than in those with normal muscle mass. The age-adjusted odds for incident low muscle mass comparing the lowest and highest FT3/FT4 increased in men. Conclusions: FT3/FT4 was positively correlated with ASM and ASMI in both men and women. Therefore, FT3/FT4 may be a parameter indicative of low muscle mass in euthyroid men with T2DM.

Protective Effect of Irbesartan by Inhibiting ANGPTL2 Expression in Diabetic Kidney Disease.

Angiopoietin-like protein 2 (ANGPTL2) stimulates inflammation and is important in the pathogenesis of diabetic kidney disease (DKD). Irbesartan is helpful in reducing diabetes-induced renal damage. In this study, the effects of irbesartan on DKD and its renal protective role involving ANGPTL2 in DKD rats were examined. Wistar rats were divided into normal, DKD, and DKD + irbesartan groups. The DKD + irbesartan group was treated once daily for 8 weeks with 50 mg/kg irbesartan via intragastric gavage. The 24-h urinary albumin was determined each week, renal pathological changes were observed, and expression of ANGPTL2 and nuclear factor-kappa B (NF-κB) in rat renal tissue was assessed by immunohistochemistry. Mouse podocytes cultured in a high concentration of glucose were classified into four groups based on the irbesartan concentrations (0, 25, 50, and 75 ºg/mL). Expression of ANGPTL2 and phosphorylated IκB-α was assessed by Western blotting. The mRNA levels of ANGPTL2 and monocyte chemotactic protein 1 (MCP-1) were assessed by real-time polymerase chain reaction. The DKD rats displayed proteinuria, podocyte injury, and increased ANGPTL2 and NF-κB expression. All were relieved by irbesartan treatment. In podocytes cultured in elevated glucose, ANGPTL2 and phosphorylated IκB-α were overexpressed at the protein level, and ANGPTL2 and MCP-1 were highly expressed at the mRNA level. Irbesartan down-regulated ANGPTL2 and phosphorylated IκB-αexpression at the protein level and inhibited ANGPTL2 and MCP-1 expression at the mRNA level. The ameliorative effects of irbesartan against DKD involves podocyte protection and suppression of ANGPTL2.

Feasibility Analysis of Cell-Free DNA Derived from Plasma of Lung Cancer Patients for Next-Generation Sequencing.

Purpose: The quality of specimens directly affects the experimental results. The stability and structural integrity of nucleic acids in samples have a decisive influence on high-throughput sequencing results. Next-generation sequencing (NGS) provides the most comprehensive criteria for evaluating the specimen quality. To test the quality of cell-free DNA (cfDNA) from lung cancer plasma samples stored in our biobank, we conducted a study to evaluate the quality in terms of the genetic level. Methods: A total of 189 peripheral blood samples were collected from patients from patients with EGFR-positive nonsmall cell lung cancer who were seen and treated in Jilin Provincial Cancer Hospital from August 2012 to March 2018. Twelve milliliters of peripheral blood samples were collected and centrifuged at 4°C, 2000 rpm for 15 minutes. Plasma samples were dispensed into cryotubes and stored at -80°C. Plasma cfDNA was extracted by a DNA extraction kit (Qiagen) and the DNA concentration was detected by a Qubit 3.0 fluorometer. Results: The total volume of cfDNA extraction at baseline was 50 µL, the median concentration according to Qubit was 0.633 ng/µL, the range was 0.331-6.09 ng/µL, and the median total DNA was 34.25 ng, ranging from 20.35 to 304.5 ng. The median value of the Qubit concentration in advanced plasma samples was 0.838 ng/µL, ranging from 0.24 to 21.9 ng/µL, and median total DNA was 41.9 ng, ranging from 12.0 to 1095.0 ng. Based on the aforementioned quality assessment factors, 4 of 189 frozen lung cancer baseline plasma samples were not included in further analyses, and for the remaining 185 cases of cfDNA >20 ng, the pass rate was 97.9%. In 143 frozen lung cancer advanced stage plasma samples, 133 cases of cfDNA >20 ng, the pass rate was 93%. Conclusion: Frozen lung cancer plasma samples stored in the biobank for 1-6 years at -80°C under certain conditions still retain a high level of cfDNA, which is suitable for NGS detection.

CX3CR1 contributes to streptozotocin-induced mechanical allodynia in the mouse spinal cord.

Patients with diabetic peripheral neuropathy experience debilitating pain that significantly affects their quality of life (Abbott et al., 2011), by causing sleeping disorders, anxiety, and depression (Dermanovic Dobrota et al., 2014). The primary clinical manifestation of painful diabetic neuropathy (PDN) is mechanical hypersensitivity, also known as mechanical allodynia (MA) (Callaghan et al., 2012). MA's underlying mechanism remains poorly understood, and so far, based on symptomatic treatment, it has no effective therapy (Moore et al., 2014).

Spinal P2X7R contributes to streptozotocin-induced mechanical allodynia in mice.

Painful diabetic neuropathy (PDN) is a diabetes mellitus complication. Unfortunately, the mechanisms underlying PDN are still poorly understood. Adenosine triphosphate (ATP)-gated P2X7 receptor (P2X7R) plays a pivotal role in non-diabetic neuropathic pain, but little is known about its effects on streptozotocin (STZ)-induced peripheral neuropathy. Here, we explored whether spinal cord P2X7R was correlated with the generation of mechanical allodynia (MA) in STZ-induced type 1 diabetic neuropathy in mice. MA was assessed by measuring paw withdrawal thresholds and western blotting. Immunohistochemistry was applied to analyze the protein expression levels and localization of P2X7R. STZ-induced mice expressed increased P2X7R in the dorsal horn of the lumbar spinal cord during MA. Mice injected intrathecally with a selective antagonist of P2X7R and P2X7R knockout (KO) mice both presented attenuated progression of MA. Double-immunofluorescent labeling demonstrated that P2X7R-positive cells were mostly co-expressed with Iba1 (a microglia marker). Our results suggest that P2X7R plays an important role in the development of MA and could be used as a cellular target for treating PDN.

Simultaneous quantitative analysis of polyethylene glycol (PEG), PEGylated paclitaxel and paclitaxel in rats by MS/MSALL technique with hybrid quadrupole time-of-flight mass spectrometry.

PEGylation is practically one of most important modifications of drugs including small molecules, peptides and proteins, which has been proven to dramatically improve physicochemical properties and pharmacokinetic behavior of the PEGylated drugs. However, it is a challenge currently to quantitatively analyze PEG and PEGylated drugs by various analytical methods, even mass spectrometry because of multiple parent ion distribution of PEG caused by its polydispersity of molecular weight. Here we developed a robust method with MS/MSALL technique using electrospray ionization (ESI) source coupled high resolution Quadrupole Time-of-Flight (Q-TOF) mass spectrometry for the quantification of PEG2K-Paclitaxel (PEG-PTX) and its two metabolites, PEG and Paclitaxel (PTX). The analysis was performed on a 300SB-C18 column with acetonitrile and 0.1% formic acid as the mobile phase. Samples were simply prepared by protein precipitation in a small quantity of plasma (50µL). Calibration curve was linear within the range of 50.0-4000ng/mL for PEG and PEG-PTX and 1.0-1000ng/mL for PTX. The intra- and inter-day precisions were 3.2-6.9% and 3.1-6.9% for PEG, 4.1-7.8% and 4.0-9.9% for PEG-PTX, and 3.3-4.8% and 3.1-6.9% for PTX, respectively. The recoveries were greater than 90% with low matrix effects. Afterwards, the newly developed method was successfully applied to support a preclinical pharmacokinetic study in six rats after single intravenous injection of PEG-PTX (51.7mg/kg).

Systemic dexmedetomidine attenuates mechanical allodynia through extracellular sign db type 2 diabetic mice.

Painful diabetic neuropathy (PDN) is a common complication of diabetes mellitus. However, the treatment for PDN is limited in clinical practice. In the present study, we investigated the effect of systemic administration dexmedetomidine (DEX), a selective alpha 2 adrenergic receptor (α2AR) agonist, on mechanical allodynia and its underlying mechanism in db/db mice, an animal model of type 2 diabetes mellitus. Our data demonstrated that db/db mice develop mechanical allodynia at the early stage of diabetes. During the period of mechanical allodynia, we detected increased release of norepinephrine (NE) and decreased levels of α2A-Adrenoceptors in db/db mice. Immunohistochemistry showed that the α2A-Adrenoceptor is predominantly expressed in neurons in the spinal cord. Acute injection of dexmedetomidine significantly decreased mechanical allodynia, which was blocked by its selective antagonist BRL44408. Furthermore, the upregulation of pERK1 and pERK2 in db/db mice were attenuated by preadministration of dexmedetomidine. We provide the first evidence that the functional alternation of spinal noradrenergic system might underlie exaggerated nociception in PDN. Systemic dexmedetomidine inhibits the mechanical allodynia which is related to ERK signaling pathway in type 2 diabetes, implying that the α2-Adrenoceptor might be a potential therapeutic strategy for PDN.

Synthesis of new amino sugar derivatives from keto-sugars of D-xylose.

Several amino sugars and imino sugar derivatives were synthesized from keto-sugars of D-xylose through a series of reactions such as the Henry reaction, hydrogenation reactions, and nucleophilic addition reactions or substitution reactions. Thiazine derivative 15 was obtained by the reaction of the keto-sugar with NH(2)CSNH(2). Higher carbon sugar 16 was accidentally prepared at room temperature from the keto-sugar in the presence of NH(2)CONH(2). The structures of the compounds were confirmed by spectral analysis. The absolute configurations of all asymmetric carbon atoms of 6 and 8 were confirmed by X-ray crystallographic analysis.

Simultaneous determination of carboprost methylate and its active metabolite carboprost in dog plasma by liquid chromatography-tandem mass spectrometry with positive/negative ion-switching electrospray ionization and its application to a pharmacokinetic study.

A liquid chromatography-tandem mass spectrometric (LC-MS/MS) method using positive/negative electrospray ionization (ESI) switching for the simultaneous quantitation of carboprost methylate and carboprost in dog plasma has been developed and validated. After screening, the esterase inhibitor, dichlorvos was added to the whole blood at a ratio of 1:99 (v/v) to stabilize carboprost methylate during blood collection, sample storage and LLE. Indomethacin was added to plasma to inhibit prostaglandins synthesis after sampling. After liquid-liquid extraction of 500µL plasma with ethyl ether-dichloromethane (75:25, v/v), analytes and internal standard (IS), alprostadil-d4, were chromatographed on a CAPCELL PAK Phenyl column (150×2.0mm, 5µm) using acetonitrile-5mM ammonium acetate as mobile phase. Carboprost methylate was detected by positive ion electrospray ionization followed by multiple reaction monitoring (MRM) of the transition at m/z 400.5→329.3; the carboprost and IS were detected by negative ion electrospray ionization followed by MRM of the transitions at m/z 367.2→323.2, and 357.1→321.2, respectively. The method was linear for both analytes in the concentration range 0.05-30ng/mL with intra- and inter-day precisions (as relative standard deviation) of ≤6.75% and accuracy (as relative error) of ≤7.21% and limit of detection (LOD) values were 10 and 20pg/mL, respectively. The method was successfully applied to a pharmacokinetic study of the analytes in beagle dogs after intravaginal administration of a suppository containing 0.5mg carboprost methylate.

Earth's free oscillations excited by the 26 December 2004 Sumatra-Andaman earthquake.

At periods greater than 1000 seconds, Earth's seismic free oscillations have anomalously large amplitude when referenced to the Harvard Centroid Moment Tensor fault mechanism, which is estimated from 300- to 500-second surface waves. By using more realistic rupture models on a steeper fault derived from seismic body and surface waves, we approximated free oscillation amplitudes with a seismic moment (6.5 x 10(22) Newton.meters) that corresponds to a moment magnitude of 9.15. With a rupture duration of 600 seconds, the fault-rupture models represent seismic observations adequately but underpredict geodetic displacements that argue for slow fault motion beneath the Nicobar and Andaman islands.