BACKGROUND:As a degradable scaffold material for bone tissue engineering,lactic acid is widely used in tissue regeneration and repair research,and plays an important role in promoting tissue healing,new bone formation and angiogenesis. OBJECTIVE:To observe the effect of lactic acid degradation products on osteoclasts and to investigate the effects of lactic-interfered osteoclast conditioned medium on the proliferation,migration and tube-forming capacity of human umbilical vein endothelial cells. METHODS:(1)The mouse monocyte macrophage cell line RAW264.7 at logarithmic growth period was selected,and adherent cells were cultured in the osteoclast induction medium(DMEM medium with nuclear factor-κB receptor-activating factor ligand and 10%fetal bovine serum)containing different concentrations of lactic acid(0,5,10,20 mmol/L).After 5 days of culture,tartrate-resistant acid phosphatase staining and cytoskeletal fibrillar actin staining were conducted.After 24 hours of culture,RT-PCR was used to detect the mRNA expression of tartrate-resistant acid phosphatase 5.(2)RAW264.7 cells at logarithmic growth period were selected and adherent cells were divided into two groups.Control group was cultured in the osteoclast induction medium,while experimental group was cultured in the osteoclast induction medium containing 10 mmol/L lactic acid.After 5 days of culture,the medium in each group was removed and the cells in the two groups were cultured in the serum-free DMEM medium for another 24 hours.Cell supernatant was then collected and used as the conditioned medium after mixed with an equal volume of DMEM medium containing 10%fetal bovine serum.Human umbilical vein endothelial cells at the logarithmic growth phase were taken and separately co-cultured with the conditioned medium of the control and experimental groups.The proliferation,migration and tube-forming ability of human umbilical vein endothelial cells were observed by cell counting kit-8 assay,migration assay,scratch assay and tube-forming assay.The mRNA and protein expression of angiogenesis-related genes and proteins were observed by RT-PCR and western blot. RESULTS AND CONCLUSION:Tartrate-resistant acid phosphatase staining and cytoskeletal fibrillar actin staining showed that 5 and 10 mmol/L lactic acid promoted osteoclastic differentiation of RAW264.7 cells and the promoting effect of 10 mmol/L lactate was more significant.RT-PCR results showed that the expression of tartrate-resistant acid phosphatase-5 mRNA of osteoclast-related genes was the highest when the lactic acid concentration was 5,10,and 20 mmol/L(P<0.05),especially 10 mmol/L.Compared with the control group,the proliferation,migration and tube-forming abilities of human umbilical vein endothelial cells were significantly increased in the experimental group(P<0.05).Compared with the control group,the expression levels of vascular endothelial growth factor and angiogenin 1 mRNA and protein were increased in the experimental group(P<0.05).To conclude,lactate-induced osteoclast conditioned medium could promote the angiogenesis of endothelial cells,and the mechanism may be related to the promotion of the expression of vascular endothelial growth factor and angiogenin 1.

Objective To explore the effects of Shuanglu Tongnao Formula on neuroinflammation in ischemic stroke (IS) rats via the P2X purinoceptor 7 receptor (P2X7R)/NLR family pyrin domain-containing 3 (NLRP3) pathway. Methods The rats were divided into five groups: the IS group, control group, Shuanglu Tongnao Formula group, P2X7R inhibitor brilliant blue G (BBG) group, and Shuanglu Tongnao Formula combined with P2X7R activator adenosine triphosphate (ATP) group, with 18 rats in each group. Except for the control group, rats in all other groups were used to construct an IS model using the suture method. After successful modeling, the drug was given once a day for 2 weeks. Neurological function scores and cerebral infarction volume ratios were measured in rats. Pathological examination of the ischemic penumbra brain tissue was performed. Immunofluorescence staining was used to quantify the proportions of microglia co-expressing both inducible nitric oxide synthase (iNOS) and ionized calcium-binding adapter molecule 1 (Iba1), as well as arginase 1 (Arg1) and Iba1, in the ischemic penumbra brain tissue. ELISA was used to detect tumor necrosis factor-alpha (TNF-α), transforming growth factor-beta (TGF-β), interleukin 6 (IL-6) and IL-10 in the ischemic penumbra brain tissue. Western blotting was used to measure P2X7R, NLRP3, and IL-1β proteins in the ischemic penumbra brain tissue. Results Compared with the control group, the IS group showed disordered neuronal arrangement, nuclear condensation, and obvious infiltration of inflammatory cells in the ischemic penumbra; significantly elevated neurological function scores, cerebral infarction volume ratios, proportions of microglia co-expressing iNOS and Iba1, and levels of TNF-α, IL-6, and P2X7R, NLRP3, IL-1β proteins; along with reduced proportions of microglia co-expressing Arg1 and Iba1 and levels of TGF-β and IL-10. Compared with the IS group, the Zhuang medicine Shuanglu Tongnao Formula and BBG groups demonstrated alleviated brain tissue damage; reduced neurological function scores, cerebral infarction volume ratios, proportions of microglia co-expressing iNOS and Iba1, and levels of TNF-α, IL-6, and P2X7R, NLRP3, IL-1β proteins; along with increased proportions of microglia co-expressing Arg1 and Iba1 and levels of TGF-β and IL-10. ATP reversed the effects of Zhuang medicine Shuanglu Tongnao Formula on microglial polarization and neuroinflammation in IS rats. Conclusion Zhuang medicine Shuanglu Tongnao Formula may promote the transformation of microglia from M1 type to M2 type by inhibiting the P2X7R/NLRP3 pathway, thereby improving neuroinflammation in IS rats.
Animals ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Receptors, Purinergic P2X7/metabolism* ; Male ; Drugs, Chinese Herbal/pharmacology* ; Rats ; Ischemic Stroke/pathology* ; Rats, Sprague-Dawley ; Disease Models, Animal ; Signal Transduction/drug effects* ; Neuroinflammatory Diseases/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Nitric Oxide Synthase Type II/metabolism* ; Interleukin-10/metabolism* ; Brain Ischemia/drug therapy* ; Microglia/metabolism*

The PCR-amplified severe fever with thrombocytopenia syndrome virus(SFTSV)Gn-DⅢ-Ⅲ gene was inserted into the pET-30a(+)prokaryotic expression vector to generate the re-combinant plasmid pET-SFTSV-Gn-D Ⅲ-Ⅲ.The plasmid was transformed into E.coli BL21(DE3)for Gn-DⅢ-m protein expression and the expression conditions were optimized.The Gn-DⅢ-Ⅲ protein purified with Ni-NTA column affinity chromatography was applied as the captured antigen to establish an indirect ELISA method for the detection of SFTSV antibody.The results demonstrated that the recombinant plasmid pET-SFTSV-Gn-D Ⅲ-Ⅲ was successfully constructed as identified by PCR and sequencing.The recombinant protein SFTSV Gn-D m-Ⅲ was soluble ex-pression in E.coli under the optimal induction conditions of 0.4 mmol/L IPTG at 25 ℃ for 4 h,and the protein purity was 91.77％after purification by Ni-NTA column.The optimal reaction con-ditions for the indirect ELISA of SFTSV antibody were as follows:coating antigen concentration(5 μg/mL),primary antibody(incubation at 37 ℃ for 1.5 h),and secondary antibody(diluted 1:10 000 and incubated at 37 ℃ for 1 h).The established method had no cross-reactivity with Rift Valley fever virus(RVFV),Ebola virus(EBOV),and tick-borne encephalitis virus(TBEV)posi-tive sera.The method had a high sensitivity,with P/N＞2.1 for SFTSV-positive sera diluted to 81920.Coefficients of variation for intra-and inter-batch reactions were less than 10％.Detection of four SFTSV-infected human clinical serum samples showed the serum samples from patients in re-mission were tested as positive(P/N＞2.1),while serum samples from patients with multiple or-gan failure were detected as negative(P/N＜2.1).The results indicated that the SFTSV Gn-D Ⅲ-Ⅲ protein was successfully expressed and purified,and it was used as the coating protein to estab-lish an indirect ELISA assay for SFTSV antibody,which possesses good specificity,sensitivity and reproducibility.This method might be applied to detect human SFTSV clinical serum samples.

Objective:To evaluate the clinical application of urine sediment score (USS) in early diagnosis, etiological differentiation, staging and prognosis of acute kidney injury (AKI), and to investigate the diagnostic efficacy of independent USS and its combination with blood urea nitrogen(Bun) serum creatinine(sCr) and uric acid(UA) in AKI.Methods:From August 23 to September 28, 2023, 9 020 morning urine samples of hospitalized patients in the First Hospital of Jilin University were detected by Sysmex UF5000.A total of 3 226 ssamples with small and round cell (SRC) > 1/μl and/or CAST>1/μl were screened for microscopic examination, and 404 cases with positive renal tubular epithelial cells and/or cast were enrolled in this study. There were 218 males and 186 females, aged 59.5 (49.0, 71.0) years. The 404 cases were divided into the USS AKI group (345 cases) and the USS non-AKI group (59 cases) according to the USS results based on the microscopic findings. According to Kidney Disease: Improving Global Outcomes (KDIGO) criteria, they were divided into KDIGO criteria AKI group (63 cases) and KDIGO criteria non-AKI group (341 cases), and the AKI group was divided into renal AKI group (33 cases) and non-renal AKI group (30 cases). According to the clinical diagnosis recorded in the medical records, they were divided into clinically diagnosed AKI group (29 cases) and clinically diagnosed non-AKI group (375 cases).The χ 2 test or Fisher exact test was used to compare USS in different AKI causes and stages. Logistic regression was used to calculate the odds ratio of renal AKI and stage 3 AKI. The area under the receiver operating characteristic curve was used to evaluate the sensitivity and specificity of USS, sCr, UA and Bun alone and in combination in the diagnosis of AKI, and the best cut-off value, sensitivity and specificity in the diagnosis of AKI were calculated. P < 0.05 was considered statistically significant. Results:The USS was used to identify the etiology of KDIGO standard AKI group,and there were significant differences in USS between renal AKI group and non-renal AKI group (χ 2=11.070, P<0.001). Compared to USS=1, the odds ratio of renal AKI was 8.125 when USS≥2 (95% CI 2.208—29.901). There was a statistically significant difference in the comparison of USS between groups in each stage of the AKI staging study based on USS (χ 2=15.724, P<0.05). Compared to USS=1, the odds ratio of stage 3 AKI was 9.714 when USS≥2 (95% CI 1.145-82.390). The AUC of independent USS in the diagnosis of AKI was 0.687 (95% CI 0.618-0.757, P<0.001), the specificity was 65.7% and the sensitivity was 61.9%. The AUC of USS combined with Bun, sCr, UA in the diagnosis of AKI was 0.794 (95% CI 0.608-0.980, P<0.05), the specificity was 82.4%, and the sensitivity was 88.9%. Conclusions:There wasan increased likelihood of renal AKI or stage 3 AKI while USS≥2,and whose combination with Bun, sCr and UA will improve the diagnostic efficiency of AKI.

In order to investigate the molecular mechanism of silk/threonine protein kinase (STK)-mediated blue light response in the algal Chlamydomonas reinhardtii, phenotype identification and transcriptome analysis were conducted for C. reinhardtii STK mutant strain crstk11 (with an AphvIII box reverse insertion in stk11 gene coding region) under blue light stress. Phenotypic examination showed that under normal light (white light), there was a slight difference in growth and pigment contents between the wild-type strain CC5325 and the mutant strain crstk11. Blue light inhibited the growth and chlorophyll synthesis in crstk11 cells, but significantly promoted the accumulation of carotenoids in crstk11. Transcriptome analysis showed that 860 differential expression genes (DEG) (559 up-regulated and 301 down-regulated) were detected in mutant (STK4) vs. wild type (WT4) upon treatment under high intensity blue light for 4 days. After being treated under high intensity blue light for 8 days, a total of 1 088 DEGs (468 upregulated and 620 downregulated) were obtained in STK8 vs. WT8. KEGG enrichment analysis revealed that compared to CC5325, the crstk11 blue light responsive genes were mainly involved in catalytic activity of intracellular photosynthesis, carbon metabolism, and pigment synthesis. Among them, upregulated genes included psaA, psaB, and psaC, psbA, psbB, psbC, psbD, psbH, and L, petA, petB, and petD, as well as genes encoding ATP synthase α, β and c subunits. Downregulated genes included petF and petJ. The present study uncovered that the protein kinase CrSTK11 of C. reinhardtii may participate in the blue light response of algal cells by mediating photosynthesis as well as pigment and carbon metabolism, providing new knowledge for in-depth analysis of the mechanism of light stress resistance in the algae.
Chlamydomonas reinhardtii/genetics* ; Photosynthesis/genetics* ; Plants/metabolism* ; Protein Kinases ; Threonine/metabolism* ; Carbon/metabolism* ; Serine/metabolism*

Objective:To summarize the clinical features of special portal hypertension-Abernethy malformation reported at home and abroad.Methods:The relevant literature on Abernethy malformation published at home and abroad from January 1989 to August 2021 was collected. Patients'clinical features, imaging and laboratory test results, diagnosis, treatment, and prognosis were analyzed.Results:A total of 380 cases were included from 60 and 202 domestic and foreign literatures. Among them, there were 200 cases of type I, with 86 males and 114 females, and the average age was (17.08±19.42) years, while there were 180 cases of type II, with 106 males and 74 females, and the average was (14.85±19.60) years. The most common reason for the first visit of an Abernethy malformation patient's was gastrointestinal system symptoms such as hematemesis and hematochezia caused by portal hypertension (70.56%). Multiple malformations were present in 45.00% of type Ⅰ and 37.80% of type Ⅱ patients. The most prevalent condition was congenital heart disease (62.22%, and 73.53%). Complications related to Abernethy malformation was occurred in 127 and 105 cases with type I and type II, respectively, with liver lesions in 74.02% (94/127) and 39.05% (42/105) and hepatopulmonary syndrome of 33.07% (42/127) and 39.05% (41/105), respectively. The imaging diagnosis of type I and type II Abernethy malformations were mainly based on abdominal computed tomography (59.00%, and 76.11%). Liver pathology was performed in 27.10% of patients. Blood ammonia increased by 89.06% and 87.50%, and AFP increased by 29.63% and 40.00% in laboratory findings. 9.76% (8/82) and 6.92% (9/130) died, while 84.15% (61/82) and 88.46% (115/130) had improved conditions after medical conservative, or surgical treatment.Conclusion:Abernethy malformation is a rare disease in which congenital portal vein development abnormalities lead to significant portal hypertension and portasystemtic shunt. Patients often seek medical treatment for gastrointestinal bleeding and abdominal pain. Type Ⅰ is more common in women, often associated with multiple malformations, and prone to secondary intrahepatic tumors. Liver transplantation is the main treatment method. Type Ⅱ is more prevalent in males, and shunt vessel occlusion is the first treatment choice. Overall, type Ⅱ has a better therapeutic impact than type Ⅰ.

The unicellular green alga Haematococcus pluvialis is the best source of natural astaxanthin (AST) in the world due to its high content under stress conditions. Although high light (HL) can effectively induce AST biosynthesis, the specific mechanisms of light signal perception and transduction are unclear. In the current study, we used transcriptomic data of normal (N), high white light (W), and high blue light (B) to study the mechanisms of light inducing AST accumulation from the point of photoreceptors. The original data of 4.0 G, 3.8 G, and 3.6 G for N, W, and B were obtained, respectively, by the Illumina Hi-seq 2000 sequencing technology. Totally, 51 954 unigenes (at least 200 bp in length) were generated, of which, 20 537 unigenes were annotated into at least one database (NR, NT, KO, SwissProt, Pfam, GO, or KOG). There were 1 255 DEGs in the W vs N, 1 494 DEGs in the B vs N, and 1 008 DEGs in the both W vs N and B vs N. KEGG enrichment analysis revealed that photosynthesis, oxidative phosphorylation, carotenoid biosynthesis, fatty acids biosynthesis, DNA replication, nitrogen metabolism, and carbon metabolism were the significantly enriched pathways. Moreover, a large number of genes encoding photoreceptors and predicted interacting proteins were predicted in Haematococcus transcriptome data. These genes showed significant differences at transcriptional expression levels. In addition, 15 related DEGs were selected and tested by qRT-PCR and the results were significantly correlated with the transcriptome data. The above results indicate that the signal transduction pathway of "light signal - photoreceptors - interaction proteins - (interaction proteins - transcription factor/transcriptional regulator) - gene expression - AST accumulation" might play important roles in the regulation process, and provide reference for further understanding the transcriptional regulation mechanisms of AST accumulation under HL stress.
Chlorophyta/genetics* ; Gene Expression Profiling ; Signal Transduction/genetics* ; Transcriptome/genetics* ; Xanthophylls

Objective:To investigate the clinical effects of anterior cervical discectomy and fusion combined with zero-profile anchored spacer (ROI-C) application on cervical spondylosis.Methods:Eighty-nine patients, consisting of 58 males and 31 females, who received anterior cervical discectomy and fusion combined with zero-profile anchored spacer (ROI-C) application in First People's Hospital of Guiyang from May 2015 to May 2018 were retrospectively analyzed. A total of 138 segments were fused. Operative time, intraoperative blood loss, postoperative complications and their incidence were recorded. The Japanese Orthopaedic Association score, Visual Analogue Scale score, and Neck Disability Index score were used to evaluate clinical effects of anterior cervical discectomy and fusion combined with ROI-C application. Imaging examination was performed to determine cervical curvature, intervertebral space height of fused segments, interbody fusion rate and changes in adjacent segments. The stability and long-term effects of postoperative cervical spine were evaluated.Results:All 89 patients were followed up for (25.6 ± 6.8) months (range 12-36 months). In 53 patients who underwent single-segment fusion, operative time was (85.54 ± 12.53) minutes and the amount of blood loss was (58.45 ± 10.24) mL. In 25 patients who underwent two-segment fusion, operative time was (115.57 ± 16.42) minutes and the amount of blood loss was (92.62 ± 12.44) mL. In 9 patients who underwent three-segment fusion, operative time was (148.63 ± 19.42) minutes and the amount of blood loss was (118.54 ± 11.25) mL. In 2 patients who underwent four-segment fusion, operative time and the amount of blood loss were 188 minutes and 175 mL, respectively in one patient and they were 214 minutes and 225 mL in another patient, respectively. With time went during 1 week to 12 months after surgery, Japanese Orthopaedic Association score was greatly increased, Neck Disability Index score was remarkably decreased, and Visual Analogue Scale score was also significantly decreased ( F = 11.25, 26.35, 20.26, all P < 0.05). Swallowing discomfort occurred in only 2 (2.2%) patients. No patients had incision hematoma, infection, hoarseness or choking cough. At 1 week to 12 months after surgery, cervical curvature and the height of intervertebral space of fused segments were superior to those before surgery (both P < 0.05). At 12 months after surgery, X-ray examination revealed bony fusion with no loosening and displacement of fusion cage and no obvious degeneration of adjacent segments. Conclusion:Anterior cervical discectomy and fusion combined with ROI-C application for the treatment of cervical spondylosis exhibits great therapeutic effects because it can greatly alleviate patient symptoms and improve cervical function.

Objective:To investigate the diagnosis and treatment value of intraoperative ultrasound (IOUS) in video-assisted thoracic surgery (VATS) of small solitary pulmonary nodule (SSPN).Methods:Of the 35 SSPN patients who received VATS in Tianjin Third Central Hospital from January 2016 to January 2020, the visual and touch examination (VTE) and IOUS method were used to locate pulmonary nodules during the operation. The differences between the two methods in the locating success rate and locating time were compared. The imaging findings of SSPN were classified and the sonographic characteristics of SSPN were summarized by univariate analysis.Results:The success rate of IOUS locating was 91.43%(32/35), which was higher than that of VTE 48.57%(17/35), and the difference was statistically significant (χ 2=15.310, P<0.001). The time of IOUS locating (6.23±1.93)min was shorter than that of VTE(9.98±1.56)min, and the difference was statistically significant ( t=6.940, P<0.001). The sonograms of 32 SSPN(17 malignancy and 15 benign) patients were all hypoechoic, univariate analysis showed that heterogeneous echo (χ 2=10.615, P=0.01) and unclear borderline (χ 2=10.041, P<0.001) were helpful to judge the benign or malignant. Conclusions:In video-assisted thoracic surgery, using IOUS could quickly and accurately locate and diagnose SSPN, which can shorten the operation time, improve the resection efficiency and guide the operation.

Objective:To investigate the awareness of digestive system injury caused by coronavirus disease 2019 (COVID-19) in gastroenterologists.Methods:From February 21 to 23 in 2020, the electronic questionnaire was used to learn about the condition of the basic knowledge of COVID-19 and awareness of digestive system injury caused by COVID-19 among the gastroenterologists across the country. Chi-square test was used for statistical analysis.Results:A total of 2 216 gastroenterologists from 31 provinces, autonomous regions and municipalities nationwide completed the survey. 99.7% (2 209/2 216) of gastroenterologists stated that they had read the COVID-19 diagnosis and treatment guidelines. The percentages of physicians who knew clearly about the diagnostic criteria of suspected and confirmed cases of COVID-19 was 34.9% (774/2 216) and 39.4% (874/2 216), respectively. The percentages of physician who gave the right answers of COVID-19 detectable methods and pulmonary imaging was 68.4% (1 516/2 216) and 71.6% (1 586/2 216), respectively. The percentages of correct answers of digestive system injury caused by COVID-19 were physicians and chief physicians was 30.9% (134/433) in residents, 33.9% (234/691) attending physicians, 32.4% (213/657) associated chief physicians and 34.9% (152/435) chief physicians, respectively, however there were no statistically significant differences among physicians with different professional titles ( χ2=6.60, P> 0.05). 95.6% (2 119/2 216) of gastroenterologists believed that probiotics could effectively improve bowel function, and 94.0% (2 082/2 216) of gastroenterologists considered that enteral nutrition support could improve patients’ prognosis. Conclusions:The knowledge and dynamic progress of the digestive system injury caused by COVID-19 are still insufficiently grasped by gastroenterologists in China. So it is necessary to carry out systematic and pertinent training programmes for them.