AIM: To explore the effect of adiponectin(ADPN)on angiogenesis of human retinal microvascular endothelial cells(hRMECs)in high glucose(HG)environment and role of NOD-like receptor family pyrin domain containing 3(NLRP3)inflammasome.METHODS: The hRMECs were divided into six groups, including control group(without treatment), HG group: incubated with D-glucose, ADPN group: pretreatment with ADPN and then incubated with D-glucose, CY-09 group: pretreatment with CY-09(an NLRP3 inhibitor)and then incubated with D-glucose, Nigericin group: pretreatment with nigericin(an NLRP3 activator)and then incubated with D-glucose, Nigericin+ADPN group: pretreatment with nigericin and ADPN and then incubated with D-glucose. NLRP3 level was detected using Western blot analysis. hRMECs migration was measured using scratch wound healing assay. The tube formation of hRMECs was detected using Matrigel.RESULTS: The NLRP3 expression in hRMECs cultured in an HG environment was significantly increased(P<0.01), while ADPN and CY-09 reduced the elevated NLRP3(both P<0.05 vs HG group). Nigericin significantly increased NLRP3 levels(P<0.01 vs control group)which was reversed by ADPN(P=0.032 vs Nigericin group). hRMECs migration ability(P<0.001), and total master segments length and number of meshes increased in HG group(P<0.001)while decreased in ADPN and CY-09 groups(all P<0.01 vs HG group). The hRMECs migration ability and tube formation(total master segments length and number of meshes)in HG environment were significantly increased by nigericin(P=0.003), while ADPN inversed the change. CONCLUSION: ADPN alleviates the migration and angiogenesis of hRMECs under HG conditions.

OBJECTIVE To investigate the clinical characteristics of euglycemic diabetic ketoacidosis （euDKA） induced by empagliflozin， and provide references for safe medication. METHODS Case reports of euDKA induced by empagliflozin were retrieved and analyzed from PubMed， Web of Science， CNKI， and Wanfang Data. The Naranjo’s scale was adopted to assess the causal relationship between empagliflozin and euDKA. RESULTS A total of 29 patients were enrolled， including 16 males and 13 females，with a median age of 58 years. All patients were diagnosed with type 2 diabetes mellitus； 22 patients had at least one comorbidity. The median time to euDKA onset was 60 days after administration， and 7 cases occurred within 2 weeks of treatment. Perioperative periods，fasting，and low-carbohydrate or ketogenic diets were the main predisposing factors. Common presentations included nausea，vomiting， fatigue， tachypnea，and tachycardia. The causal relationship between empagliflozin and euDKA was assessed as “probable” in all cases. According to adverse reaction severity grading， 13 cases were grade 3 （severe） and 16 cases were grade 4 （life-threatening）. All patients discontinued empagliflozin and recovered after corresponding treatment. CONCLUSIONS euDKA is a serious adverse reaction to empagliflozin， which mostly occurs early in treatment. Perioperative periods， fasting， and ketogenic diets are prone to induce euDKA. Elderly patients and those with comorbidities are at higher risk. Risk assessment should be conducted before clinical administration； blood ketones and acid-base monitoring should be strengthened in the early medication period； withhold the drug before elective surgery， avoid prolonged fasting；upon diagnosis， discontinue empagliflozin and initiate intravenous insulin， provide fluid resuscitation promptly， and use other treatments to improve patient prognosis.

ObjectiveTo study the effect of Taikong Yangxin Pills on the metabolism of simulated weightless rats based on metabolomics and discuss the metabolism mechanism. MethodsIn the simulated space capsule environment on the ground， the rat model of simulated weightlessness was established by the tail suspension method. Rats were randomly grouped as follows： out-of-capsule control， in-capsule control， model， and high （3.0 g·kg^-1） and low （1.5 g·kg^-1） doses of Taikong Yangxin Pills， and they were administrated with corresponding drugs by gavage for 28 days. The serum levels of endogenous metabolites in rats were determined by ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry （UPLC-Q-TOF/MS）. The obtained data were processed by principal component analysis （PCA） and orthogonal partial least squares-discriminant analysis （OPLS-DA） to screen for differential metabolites and potential biomarkers. MetaboAnalyst 5.0 was used for pathway enrichment analysis to explain the metabolic regulation mechanism of the drug. ResultsCompared with the out-of-capsule control group， the in-capsule control group showed elevated levels of thirteen metabolites， including 14-hydroxyhexadecanoic acid， linoleic acid， and α-linolenic acid （P<0.05）， which suggested that the space capsule environment mainly affected the metabolism of α-linolenic acid and linoleic acid in the rats. Compared with the in-capsule control group， the model group showed lowered levels of fourteen metabolites， including 4-imidazolone-5-propionic acid， isocitric acid/citric acid， and L-tyrosine （P<0.05）， which were recovered after the treatment with Taikong Yangxin pills （P<0.05）. The pathway enrichment analysis revealed that weightlessness induced by tail suspension and drug intervention mainly involved the phenylalanine， tyrosine， and tryptophan biosynthesis， tyrosine metabolism， histidine metabolism， and citric acid cycle. ConclusionThe simulated space capsule environment and simulated weightlessness induced by tail suspension can both affect the metabolism level of rats. Taikong Yangxin pills can ameliorate the metabolic abnormality in the rat model of weightlessness by regulating various amino acids and energy metabolism-related pathways.

ObjectiveTo establish the fingerprints of 15 batches of Hengzhi Kechuan capsules， to quantitatively analyze 10 index components， and to evaluate the quality uniformity of samples from different batches. MethodsThe fingerprints and quantitative analysis of Hengzhi Kechuan capsules were established by a combination method of high performance liquid chromatography coupled with diode array detector and charged aerosol detector（HPLC-DAD-CAD）， adenosine， guanosine， vanillic acid， safflomin A， agarotetrol， naringin， hesperidin， militarine， ginsenoside Rb₁， and glycyrrhizic acid were selected as quality attribute indexes. A total of 15 batches of Hengzhi Kechuan capsules from 2022 to 2024（3 boxes per batch） were qualitatively and quantitatively analyzed， and the quality uniformity level of the manufacturers was characterized by parameters of intra-batch consistency（P_A） and inter-batch consistency（P_B）. The homogeneity and difference of quality attribute indexes of samples from different years were analyzed by heatmap clustering analysis. ResultsHPLC fingerprints and quantitative method of Hengzhi Kechuan capsules were established， and the methods could be used for qualitative and quantitative analysis of this preparation， which was found to be stable and reliable by method validation. The similarity of fingerprints of 15 batches of samples was 0.887-0.975， a total of 13 common peaks were calibrated， and 10 common peaks were designated， all of which were quality attribute index components. The results of quantitative analysis showed that the contents of the above 10 ingredients in the samples were 0.038-0.078， 0.115-0.251， 0.007-0.018， 0.291-0.673， 0.122-0.257， 0.887-1.905， 1.841-3.364， 1.412-2.450， 2.207-3.112， 0.650-1.161， respectively. And the contents of ginsenoside Rb₁ and glycyrrhizic acid met the limit requirements in the 2020 edition of Chinese Pharmacopoeia. For the samples from 15 batches， the P_A values of the 10 index components were all <10%， indicating good intra-batch homogeneity， and the P_B values ranged from 33.86% to 92.97%， suggesting that the inter-batch homogeneity was poor. Heatmap clustering analysis showed that the samples from different years were clustered into separate categories， and adenosine， guanosine， safflomin A， naringin， hesperidin and agarotetrol were the main differential components. ConclusionThe intra-annual quality uniformity of Hengzhi Kechuan capsules is good and the inter-annual quality uniformity is insufficient， which may be related to the quality difference of Pinellinae Rhizoma Praeparatum， Carthami Flos， Citri Sarcodactylis Fructus， Citri Reticulatae Pericarpium， Aquilariae Lignum Resinatum， Citri Fructus， etc. In this study， the fingerprint and multi-indicator determination method of Hengzhi Kechuan capsules was established， which can be used for more accurate and efficient quality control and standardization enhancement.

ObjectiveTo explore the characteristics of ecological niche and interspecific relationships of mosquitoes in different habitats in Dongcheng District, Beijing, and to provide a basis for mosquito ecological monitoring, control and the development or optimization of prevention and control strategies for related mosquito-borne diseases. MethodsFrom May to October 2023, the ecological monitoring in residential areas, parks, tourist attractions and medical institutions in Dongcheng District of Beijing was carried out using the carbon dioxide (CO₂) mosquito trapping method, and the ecological niche characteristics and interspecific relationships of mosquitoes in different habitats were analyzed using Levins ecological niche breadth index, Pinaka ecological niche overlap index and ecological niche similarity coefficients. ResultsThe temporal ecological niche of Culex pipiens pallens (10.62) was higher than that of Aedes albopictus (8.29) in different habitats in Dongcheng District of Beijing, and the temporal ecological niche overlap index of the two mosquitoes was as high as 0.87. The ecological niche breadth of Culex pipiens pallens was higher than that of Aedes albopictus in different monitoring habitats, and the order of the ecological niche breadth of Culex pipiens pallens in different monitoring habitats was, from high to low, as follows: residential areas (11.09) > tourist attractions (10.25) > medical institutions (9.15) > parks (9.07), while the ecological niche breadth of Aedes albopictus in different habitats was, in descending order, residential areas (8.56) > medical institutions (7.68) > parks (7.44) > tourist attractions (5.73). The results of niche overlap analysis showed that the overlap index between Culex pipiens pallens and Aedes albopictus was the largest in residential areas (0.86), as for in other habitats, which was, in descending order, parks (0.81) > medical institutions (0.68) > tourist attractions (0.60). Besides, the ecological similarity coefficients further verified that similarity coefficients, between the two mosquito species, were highest in residential areas (0.712), lowest in tourist attractions (0.497), and which were 0.675 in parks and 0.598 in medical institutions, respectively. ConclusionIn different monitoring habitats in Dongcheng District of Beijing, Culex albopictus pallens demonstrates a stronger spatio-temporal resource utilization ability than Aedes albopictus, and the two species exhibit more similar spatio-temporal resource utilization patterns in residential areas. Corresponding control strategies targeting the characteristics of ecological niches and interspecific relationships of these two mosquito species in different habitats should be developed to enhance the prevention and control effect.

The wrist joint is a highly mobile functional joint. Wrist conditions including traumatic and degenerative arthritis, rheumatoid arthritis, and giant cell tumors of the distal radius, cause significant pain and mobility impairment. In joint surgery, the decision to use joint prostheses to reconstruct joint function is greatly influenced by the characteristics of the prosthesis (Mok et al., 2016). However, traditional implants have limitations such as shape mismatch, inadequate implant-bone interface strength which causes loosening, and poor bone ingrowth (Zhang et al., 2014).
Humans ; Joint Prosthesis ; Wrist Joint/surgery* ; Prosthesis Design ; Arthroplasty, Replacement ; Arthritis, Rheumatoid/surgery*

Coronavirus disease 2019 (COVID-19) is a multi-system disease that can lead to various severe complications. Acute limb ischemia (ALI) has been increasingly recognized as a COVID-19-associated complication that often predicts a poor prognosis. However, the pathophysiology and molecular mechanisms underlying COVID-19-associated ALI remain poorly understood. Hypercoagulability and thrombosis are considered important mechanisms, but we also emphasize the roles of vasospasm, hypoxia, and acidosis in the pathogenesis of the disease. The angiotensin-converting enzyme 2 (ACE2) pathway, inflammation, and platelet activation may be important molecular mechanisms underlying these pathological changes induced by COVID-19. Furthermore, we discuss the hypotheses of risk factors for COVID-19-associated ALI from genetic, age, and gender perspectives based on our analysis of molecular mechanisms. Additionally, we summarize therapeutic approaches such as use of the interleukin-6 (IL-6) blocker tocilizumab, calcium channel blockers, and angiotensin-converting enzyme inhibitors, providing insights for the future treatment of coronavirus-associated limb ischemic diseases.
Humans ; COVID-19/physiopathology* ; Ischemia/etiology* ; SARS-CoV-2 ; Extremities/blood supply* ; Risk Factors ; Interleukin-6/antagonists & inhibitors* ; Acute Disease ; Angiotensin-Converting Enzyme 2

OBJECTIVES: To evaluate the protective effect of Schistosoma japonicum cystatin (rSj-Cystatin) in a mouse mode of "two-hit" sepsis. METHODS: Sixty male C57BL/6 mice randomized equally into sham-operated group, protein group, "two-hit" modeling group, and protein intervention group. In the former two groups, the mice received an intraperitoneal injection of 100 μL PBS followed by exposure of the cecum and then by intraperitoneal injection of 100 μL PBS or 25 μg rSj-Cystatin 30 min later; In the latter two groups, 100 μL PBS containing LPS (5 mg/kg) was injected intraperitoneally 24 h before cecal ligation and puncture (CLP), and 100 μL PBS or 25 μg rSj-Cystatin were injected 30 min after CLP. At 12 h after rSj-Cystatin treatment, 6 mice from each group were sacrificed for detection of TNF-α, IL-6, IL-10, TGF-β, iNOS and Arg-1 in the serum, spleen, liver, lung and kidney tissues using ELISA, for examinations of liver, lung and kidney pathologies with HE staining, and for analysis of CD3⁺CD4⁺CD25⁺Foxp3⁺ T cell percentage in the spleen using flow cytometry. The remaining mice were observed for general condition and 72-h survival. RESULTS: The 72-h survival rates in the 4 groups were 100%, 100%, 0% and 20%, respectively, showing significant differences between the latter two groups. The mouse models of "two-hit" sepsis exhibited obvious tissue pathologies and significant elevations of TNF-α and IL-6 in both the serum and tissue homogenate, which were significantly ameliorated by rSj-Cystatin treatment. Treatment with rSj-Cystatin also increased IL-10 and TGF-β levels and spleen CD3⁺CD4⁺CD25⁺Foxp3⁺ T cell percentage. The septic mouse models also showed increased iNOS levels in all the detected tissues and a decreased Arg-1 level in the kidney, and these changes were obviously improved by rSj-Cystatin treatment. CONCLUSIONS rSj-Cystatin has a protective effect against "two-hit" sepsis in mice by regulating the inflammatory microenvironment.
Animals ; Mice ; Sepsis/drug therapy* ; Male ; Schistosoma japonicum/chemistry* ; Mice, Inbred C57BL ; Cystatins/therapeutic use* ; Interleukin-10/metabolism* ; Interleukin-6/blood* ; Tumor Necrosis Factor-alpha/blood* ; Disease Models, Animal ; Transforming Growth Factor beta/metabolism*

This study investigated the specific immune response of BALB/c mice that was induced by a circular RNA (circRNA) vaccine expressing the herpes simplex virus type II (HSV-2) glycoprotein D (gD). The aim was to evaluate the immunological potential of this vaccine and lay a foundation for developing an mRNA vaccine against HSV-2. PCR and homologous recombination were employed to integrate the gD gene obtained from the pT7AMP-gD ectodomain plasmid into pUC57 to generate the recombinant plasmid pUC57-circ-gD, which was then sequenced and characterized. In vitro transcription and cyclization were performed on the template DNA to generate pUC57-circ-gD mRNA. To validate the formation of circular RNA, we cleaved the pUC57-circ-gD mRNA with RNase R and employed RT-PCR to validate the cyclization. The pUC57-circ-gD mRNA was then transfected into 293T cells. After 72 h, the cell supernatant was collected, and Western blotting was employed to measure the protein level of gD. Subsequently, the mRNA was encapsulated in lipid nanoparticles (LNPs) by microfluidic encapsulation. BALB/c mice were administrated with the encapsulated mRNA, and blood was collected from the fundus venous plexus after 21 and 35 days, and from the enucleated eyeballs after 49 days. Enzyme-linked immunosorbent assay was employed to measure the titers of antibodies, including virus-neutralizing antibodies. After 49 days, spleens were harvested and assessed for secretion of interferon-gamma (IFN-γ) by solid-phase enzyme-linked immunospot. The results showed successful construction and sequencing of the recombinant plasmid. RNase R digestion confirmed the presence of circular RNAs. Western blotting of the 293T cells transfected with the mRNA showed clear specific bands. The quality of the vaccine was tested by size exclusion chromatography-high performance liquid chromatography, which showed that the purity of the vaccine was about 90%. The mRNA-LNP showcased the particle size of 82.76 nm and an encapsulation rate of approximately 98%. Following three-dose vaccination, all immunized mice exhibited steady weight gain with 100% survival rate throughout the 28-day observation period, indicating no significant acute toxicity associated with the vaccine formulation. The immunized mice showed dose-dependent increases in serum IgG antibody titer and IFN-γ secretion by splenocytes and they were resistant to virus attacks. These findings indicate good immunogenicity and persistence of the pUC57-circ-gD mRNA vaccine, providing a reference for further studies on circRNA vaccines.
Animals ; Mice, Inbred BALB C ; RNA, Circular ; Mice ; Humans ; Herpesvirus 2, Human/genetics* ; Viral Envelope Proteins/genetics* ; Antibodies, Viral/blood* ; HEK293 Cells ; Female ; Nanoparticles ; Plasmids

This study aims to investigate the potential of oncolytic nanoparticles encapsulating Coxsackievirus A21 (CVA21) full-genome mRNA (CVA21@ONP) to resurrect CVA21 and induce apoptosis in host cells, as well as the antitumor immune effects of CVA21@ONP in immunocompetent tumor-bearing BALB/c mice. We used lipid nanoparticles (LNPs) to encapsulate CVA21 full-genome mRNA, thus preparing CVA21@ONP. The killing efficacy of CVA21@ONP was determined by the plaque assay and cell counting kit-8 (CCK-8), and the apoptosis in HT29 and CT26-iRFP cells was evaluated by flow cytometry. Mice were administrated with CVA21@ONP at high and low doses intratumorally, and the growth of tumors expressing infra-red fluorescent protein (iRFP) was monitored. Additionally, the types and changes of immune cells in the spleen were analyzed by flow cytometry. The results demonstrated that CVA21@ONP successfully resurrected CVA21 in both HT29 and U87MG cells. The plaque assay revealed robust killing effects of CVA21@ONP against both human and murine cell lines, and flow cytometry results showed increased early and late apoptotic cells. Notably, intratumoral detection revealed significantly down-regulated expression of iRFP in both high- and low-dose CVA21@ONP groups. Flow cytometry results further indicated that CVA21@ONP treatment effectively reduced the levels of immunosuppressive cells, including myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs), in the spleen, while enhancing T cell-dependent antitumor immune responses. These findings suggest that CVA21@ONP can replicate and survive extensively both in vitro and in vivo, activating the immune system of mice administrated with CVA21@ONP to target cells at the tumor site, thereby remodeling the tumor immune microenvironment and accelerating the suppression or even complete regression of tumors. The oncolytic performance of CVA21@ONP has been verified through intratumoral injection administration in this study, aimed at further exploring its therapeutic potential and promoting the development of the field of tumor treatment.
Animals ; Nanoparticles/chemistry* ; Mice ; Mice, Inbred BALB C ; Humans ; Apoptosis ; Oncolytic Viruses/genetics* ; Oncolytic Virotherapy/methods* ; Cell Line, Tumor ; RNA, Messenger/genetics* ; HT29 Cells