OBJECTIVE: To investigate the effect of stretch on long non-coding RNA taurine upregulated gene 1 (TUG1)-mediated miR-545-3p/cannbinoida receptor 2 (CNR2) pathway regulating bone regeneration in the distraction area of rats during distraction osteogenesis. METHODS: Thirty-six 10-week-old male Sprague Dawley rats were randomly divided into 3 groups ( n=12 in each group): group A (femoral fracture+injection of interfering RNA), group B (distraction osteogenesis+injection of interfering RNA), and group C (distraction osteogenesis+injection of TUG1). Groups A and B were injected with 60 μg of interfering RNA at the beginning of incubation period (immediate after operation), the beginning of distraction phase (7 days after operation), and the end of distraction phase (21 days after operation), and group C was injected with 60 μg of synthetic TUG1 in vivo interfering sequence at the same time. The general situation of rats in each group was observed during the experiment. The mineralization of fracture space or distraction area was observed by X-ray films at 21, 35, and 49 days after operation. At 49 days after operation, the samples of the distraction area were taken for HE staining to observe the mineralization, and real-time fluorescence quantitative PCR (qRT-PCR) was used to detect the expressions of osteoblast-related genes such as TUG1, miR-545-3p, CNR2, alkaline phosphatase (ALP), osteocalcin (OCN), and osteopontin (OPN). Blood samples were collected from the abdominal aorta of the rats, and the expressions of ALP and C terminal telopeptide of type Ⅰ (CTX-Ⅰ) protein were detected by ELISA assay. RESULTS: The results of X-ray film and HE staining observations showed that osteogenesis in group C was superior to groups A and B at the same time point. The results of qRT-PCR showed that the relative mRNA expressions of TUG1, CNR2, ALP, OCN, and OPN in group C were significantly higher than those in group A and group B, and the relative mRNA expression of miR-545-3p in group C was significantly lower than that in group A and group B ( P<0.05). The relative mRNA expressions of TUG1 and ALP in group B were significantly higher than those in group A, and the relative mRNA expression of miR-545-3p in group B was significantly lower than that in group A ( P<0.05). There was no significant difference in the relative mRNA expressions of CNR2, OCN, and OPN between group A and group B ( P>0.05). The results of ELISA showed that the expressions of ALP and CTX-Ⅰ protein were significantly higher in group C than in group A and group B, and in group B than in group A ( P<0.05). CONCLUSION Under the action of stretch, the expression of TUG1 in the femoral distraction area of rats increases, which promotes the expression of CNR2 by inhibiting the expression of miR-545-3P, which is helpful to the mineralization of the extension area and osteogenesis.
Animals ; MicroRNAs/genetics* ; Rats, Sprague-Dawley ; Male ; Osteogenesis, Distraction/methods* ; Rats ; RNA, Long Noncoding/metabolism* ; Osteopontin/genetics* ; Osteogenesis ; Bone Regeneration ; RNA, Small Interfering/genetics* ; Osteocalcin/genetics* ; Alkaline Phosphatase/metabolism* ; Osteoblasts/cytology* ; Signal Transduction ; Femoral Fractures/surgery*

The rapid emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants that evade immunity elicited by vaccination has posed a global challenge to the control of the coronavirus disease 2019 (COVID-19) pandemic. Therefore, developing countermeasures that broadly protect against SARS-CoV-2 and related sarbecoviruses is essential. Herein, we have developed a lipid nanoparticle (LNP)-encapsulated mRNA (mRNA-LNP) encoding the full-length Spike (S) glycoprotein of SARS-CoV-2 (termed RG001), which confers complete protection in a non-human primate model. Intramuscular immunization of two doses of RG001 in Rhesus monkey elicited robust neutralizing antibodies and cellular response against SARS-CoV-2 variants, resulting in significantly protected SARS-CoV-2-infected animals from acute lung lesions and complete inhibition of viral replication in all animals immunized with low or high doses of RG001. More importantly, the third dose of RG001 vaccination elicited effective neutralizing antibodies against current epidemic XBB and JN.1 strains and similar cellular response against SARS-CoV-2 Omicron variants (BA.1, XBB.1.16, and JN.1) were observed in immunized mice. All these results together strongly support the great potential of RG001 in preventing the infection of SARS-CoV-2 variants of concern (VOCs).

OBJECTIVE: To elucidate the role and mechanism of microRNA-145-5p (miR-145-5p) in hypoxia-induced pyroptosis of human alveolar epithelial cells. METHODS: In vitro, human alveolar epithelial cell line BEAS-2B was cultured. Cells in the logarithmic growth phase were cultured to 80% confluence and then used for the experiment. (1) BEAS-2B cells were cultured under 1% O₂ hypoxic condition, with a normoxic control group. Western blotting was employed to detect the expressions of pyroptosis marker proteins [NOD-like receptor protein 3 (NLRP3), Gasdermin D N-terminal domain (GSDMD-N), and caspase-1] in cells cultured for 24 hours. Real-time fluorescent quantitative reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the expression of miR-145-5p in cells cultured for 6 hours and 12 hours. (2) Cells were transfected with 30 nmol/L miR-145-5p mimic to overexpress miR-145-5p expression under normoxic condition or 30 nmol/L miR-145-5p inhibitor to suppress miR-145-5p expression under hypoxic condition. Control group and negative control group were respectively set up. After 24 hours of cell culture, Western blotting was used to detect the expressions of pyroptosis marker proteins and nuclear factor-E2-related factor 2 (Nrf2) in cells. Flow cytometry was applied to detect the level of reactive oxygen species (ROS) in cells. The target genes of miR-145-5p were predicted by miR target gene prediction software miRWalk and verified by Western blotting. (3) Under hypoxic condition, cells were transfected with 6.94 ng/μL silent information regulator 5 (Sirt5) overexpression plasmid or pretreated with 12.5 mmol/L N-acetyl-L-cysteine (NAC) as an ROS inhibitor. The empty plasmid group and control group were set up. After 24 hours of cell culture, Western blotting was used to detect the expressions of Sirt5, Nrf2, and pyroptosis marker proteins in cells. Flow cytometry was used to detect the level of ROS in cells. RESULTS: (1) Compared with the normoxic control group, the expression levels of pyroptosis marker proteins in the 24-hour hypoxia group was significantly increased, indicating that hypoxia could induce pyroptosis in BEAS-2B cells. The expression level of miR-145-5p in cells gradually increased with the extension of hypoxia induction time, indicating that hypoxia could cause the increase of miR-145-5p expression level. (2) The expression levels of pyroptosis marker proteins in cells of miR-145-5p mimic group significantly increased under normoxic condition as compared with the control and negative control groups [NLRP3 protein (NLRP3/β-actin): 1.58±0.07 vs. 1.00±0.01, 0.98±0.07, GSDMD-N protein (GSDMD-N/β-actin): 1.71±0.03 vs. 1.01±0.01, 0.85±0.03, caspase-1 protein (caspase-1/β-actin): 2.33±0.04 vs. 1.01±0.01, 1.05±0.04, all P < 0.05], Nrf2 protein expression level was significantly decreased (Nrf2/β-actin: 0.79±0.03 vs. 1.00±0.01, 1.03±0.04, both P < 0.05), ROS level was significantly up-regulated (fluorescence intensity: 1.74±0.03 vs. 1.00±0.01, 0.92±0.03, both P < 0.05). Under hypoxia condition, compared with control group and negative control group, the expression levels of pyroptosis marker proteins in miR-145-5p inhibitor group were significantly decreased [NLRP3 protein (NLRP3/β-actin): 0.21±0.04 vs. 1.70±0.02, 1.63±0.04; GSDMD-N protein (GSDMD-N/β-actin): 1.32±0.02 vs. 2.51±0.02, 2.72±0.03; caspase-1 protein (caspase-1/β-actin): 0.56±0.01 vs. 2.77±0.02, 3.12±0.03; all P < 0.05], Nrf2 protein expression level was significantly increased (Nrf2/β-actin: 1.57±0.04 vs. 1.22±0.01, 1.28±0.04, both P < 0.05), ROS level was significantly down-regulated (fluorescence intensity: 0.64±0.05 vs. 1.87±0.04, 1.70±0.07, both P < 0.05). The results indicated that miR-145-5p could promote cell pyrodeath. The predictive result of miRWalk showed that the 3' untranslated region (3'UTR) of Sirt5 had complementary base binding sites with miR-145-5p. The expression level of Sirt5 protein in cells of miR-145-5p mimic group was significantly lower than that of control group and negative control group under normoxic condition (Sirt5/β-actin: 0.59±0.03 vs. 1.00±0.01, 1.01±0.03, both P < 0.05), which verified that Sirt5 was the target gene of miR-145-5p. (3) The occurrence of pyrodeath could be partially reversed by transfection with Sirt5 overexpression plasmid or adding ROS inhibitor NAC into cells, and Sirt5 overexpression could also up-regulate Nrf2 expression and eliminate intracellular ROS. CONCLUSION In human alveolar epithelial cells, miR-145-5p can down-regulate Nrf2 by targeting Sirt5, thereby increasing ROS expression and inducing pyrodeath.
Humans ; MicroRNAs ; Pyroptosis ; Cell Hypoxia ; Alveolar Epithelial Cells/cytology* ; Cell Line ; NLR Family, Pyrin Domain-Containing 3 Protein ; Caspase 1/metabolism* ; Epithelial Cells/metabolism* ; Gasdermins ; Phosphate-Binding Proteins

Objective To explore the relationship between preoperative coronary angiography and postoperative acute kidney injury (AKI) in cardiac surgery. Methods The clinical data of patients who underwent coronary angiography within 30 days before cardiac surgery in the First Affiliated Hospital of Xi’an Jiaotong University from January 2015 to April 2019 were retrospectively analyzed. Univariate analysis and multivariate logistic regression analyses were used to explore the relationship between the interval from preoperative coronary angiography to cardiac surgery and postoperative AKI. Results Finally 1 112 patients were collected, including 700 males and 412 females, with a median age of 61 (55, 66) years. The incidence of postoperative AKI was 40.8% (454/1 112), of which grade 2-3 AKI accounted for 11.9%. Multivariate analysis showed that age (OR=1.049, 95%CI 1.022-1.077, P<0.001), body mass index (OR=1.065, 95%CI 1.010-1.123, P=0.020) and time interval between preoperative coronary angiography and cardiac surgery within 24 hours (OR=1.625, 95%CI 1.116-2.364, P=0.011) were independent predictors of postoperative AKI. Patients who underwent coronary angiography within 24 hours before surgery had a 10.6% higher incidence of postoperative AKI compared to those who underwent angiography ≥24 hours before surgery (P=0.004). Patients who underwent valve surgery with or without coronary artery bypass grafting (CABG) had a higher risk of AKI than those who only underwent CABG. The in-hospital stay of patients who developed AKI was 2 days longer than those without AKI. However, undergoing coronary angiography within 24 hours before cardiac surgery did not prolong the length of ICU stay or hospital stay, nor did it increase the risk of death or renal failure after the operation. Conclusion Undergoing coronary angiography within 24 hours before cardiac surgery increases the risk of postoperative AKI.

Objective:To explore the surgical technique and results of three-dimensional aortic valve anatomic repair for bicuspid aortic valve (BAV) with aortic regurgitation (AR).Methods:This is a retrospective case series study. From August 2021 to December 2023, 130 consecutive patients with BAV-AR underwent aortic valve anatomic repair at the Department of Cardiothoracic Surgery, Zhongshan Hospital, Fudan University,and the data were retrospectively analyzed. There were 115 males and 15 females, aged (38.6±11.7) years (range: 15 to 67 years). All patients received modified aortic root reconstruction, to do three-dimensional root remodeling, including the basal ring, sinus of Valsalva and sino-tubular junction simultaneously. Perioperative and follow-up data were collected and analyzed. Comparisons between groups were performed using independent samples t-test, Wilcoxon paired signed-rank test, or χ2 test. Results:No patient transferred to valve replacement during the operation. The cardiopulmonary bypass time ( M(IQR)) was 109(34) minutes (range:67 to 247 minutes), and the aortic cross-clamp time was 76(26) minutes (range: 32 to 158 minutes). Preoperative transesophageal echocardiography showed 123 patients (94.6%) presented with moderate or severe regurgitation. Immediately postoperative transesophageal echocardiography showed no regurgitation in 22 patients (16.9%), trace regurgitation in 81 patients (62.3%) and mild regurgitation in 27 patients (20.8%). Follow up was completed in all patients, with a follow-up of 5.5(9.4) months (range: 0.1 to 27.6 months). No mortality was observed during follow-up. Echocardiography was obtained in 112 patients at the latest follow-up, including no regurgitation in 4 patients (3.6%), trace regurgitation in 58 patients (51.8%), mild regurgitation in 45 patients (40.2%), moderate regurgitation in 4 patients (3.6%), and severe regurgitation in 1 patient (0.9%). Conclusion:For patients with BAV-AR who have good valve quality and no severe aortic sinus dilation, the recent outcomes of three-dimensional anatomical repair technique, focusing on overall remodeling of the aortic root, are satisfactory.

Objective:To explore the surgical technique and results of three-dimensional aortic valve anatomic repair for bicuspid aortic valve (BAV) with aortic regurgitation (AR).Methods:This is a retrospective case series study. From August 2021 to December 2023, 130 consecutive patients with BAV-AR underwent aortic valve anatomic repair at the Department of Cardiothoracic Surgery, Zhongshan Hospital, Fudan University,and the data were retrospectively analyzed. There were 115 males and 15 females, aged (38.6±11.7) years (range: 15 to 67 years). All patients received modified aortic root reconstruction, to do three-dimensional root remodeling, including the basal ring, sinus of Valsalva and sino-tubular junction simultaneously. Perioperative and follow-up data were collected and analyzed. Comparisons between groups were performed using independent samples t-test, Wilcoxon paired signed-rank test, or χ2 test. Results:No patient transferred to valve replacement during the operation. The cardiopulmonary bypass time ( M(IQR)) was 109(34) minutes (range:67 to 247 minutes), and the aortic cross-clamp time was 76(26) minutes (range: 32 to 158 minutes). Preoperative transesophageal echocardiography showed 123 patients (94.6%) presented with moderate or severe regurgitation. Immediately postoperative transesophageal echocardiography showed no regurgitation in 22 patients (16.9%), trace regurgitation in 81 patients (62.3%) and mild regurgitation in 27 patients (20.8%). Follow up was completed in all patients, with a follow-up of 5.5(9.4) months (range: 0.1 to 27.6 months). No mortality was observed during follow-up. Echocardiography was obtained in 112 patients at the latest follow-up, including no regurgitation in 4 patients (3.6%), trace regurgitation in 58 patients (51.8%), mild regurgitation in 45 patients (40.2%), moderate regurgitation in 4 patients (3.6%), and severe regurgitation in 1 patient (0.9%). Conclusion:For patients with BAV-AR who have good valve quality and no severe aortic sinus dilation, the recent outcomes of three-dimensional anatomical repair technique, focusing on overall remodeling of the aortic root, are satisfactory.

BACKGROUND:Peripheral nerves play an important role in bone metabolism.In clinical practice,the specific impact of nerve injury on bone transport technology needs further study. OBJECTIVE:To investigate the effect of tibial nerve injury on the treatment of tibial slip by single-plane osteotomy. METHODS:Thirty-two patients with tibial bone defects admitted to Tangshan Second Hospital from May 2011 to June 2022 were selected.According to the presence or absence of tibial nerve injury,patients were divided into the tibial nerve injury group(n=16)and the non-tibial nerve injury group(n=16).Both groups were treated with single-plane osteotomy and bone slip.After treatment,the patients were followed up to collect the mineralization zone healing index,external fixation index,docking point healing and needle infection.After the removal of external fixation,the bone healing and functional evaluation were evaluated by a classification of the Association for the Study and Application of the Method of Ilizarov(ASAMI). RESULTS AND CONCLUSION:(1)All 32 patients were followed up for(25.28±4.79)months.There were no significant differences in bone healing time,external fixation time,healing index and external fixation index between the two groups(P＞0.05).Needle infection occurred in two cases of the tibial nerve injury group and one case of the non-tibial nerve injury group,all of which were PALEY I,and there was no significant difference between the two groups(P＞0.05).The non-union rate of the occlusal end of the tibial nerve injury group was 31%,and that of the non-tibial nerve injury group was 13%;there was no statistical difference between the two groups(P＞0.05).The excellent and good rate of ASAMI bone healing score in the two groups was 100%;the excellent and good rate of limb score was 81%in the tibial nerve injury group and 94%in the non-tibial nerve injury group;there was no statistical difference between the two groups(P＞0.05).(2)Our research shows that tibial nerve injury has no significant effect on the mineralization speed,external fixation time,union of the occlusal end,infection of the needle tract,and the quality of bone formation in the mineralized area of the single-plane osteotomy.

ObjectiveStudy on the mechanism of Guishao Yunpi decoction in preventing and treating breast hyperplasia based on phosphatase and tensin homolog （PTEN）/ phosphatidylinositol 3-kinase （PI3K）/protein kinase B （Akt） pathway. MethodSeventy SPF-grade SD rats were randomly assigned into blank group （n=15） and modeling group （n=55）. The mammary gland hyperplasia model of liver stagnation and spleen deficiency was established by the comprehensive modeling method （hunger and satiety abnormality + tail stimulation + estradiol benzoate + progesterone）， and then 5 rats were randomly selected for model verification. The modeled rats were then randomly assigned into five groups： model group， positive control （4 mg·kg^-1·d^-1 tamoxifen） group， and high-， medium-， and low-dose （17.2， 8.6， 4.3 g·kg^-1·d^-1， respectively） Guishao Yunpi decoction groups， with 10 rats in each group. The rats in the blank group and model group were given 10 mL·kg^-1·d^-1 distilled water， and those in other groups were orally administrated with corresponding drugs. After 30 days of treatment， the general living conditions of rats were observed， and the thickness of breast tissue was measured by an ultrasonic diagnostic instrument. The open field test was carried out for behavioral evaluation. The levels of B-cell lymphoma-2 （Bcl-2） and Bcl-2-associated X （Bax） in the breast tissue were determined by enzyme-linked immunosorbent assay （ELISA）. The mRNA and protein levels of PTEN， PI3K， and Akt in the breast tissue were determined by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot， respectively. ResultCompared with those in the blank group， the rats in the model group were irritable， curled up in clusters， and showed positive behavior in the open field test. The modeling led to nipple swelling and increased the breast thickness （P<0.05）. Moreover， the modeling elevated the level of Bcl-2 and lowered that of Bax in the breast tissue （P<0.05）， down-regulated the mRNA and protein levels of PTEN， and up-regulated the mRNA and protein levels of PI3K and Akt （P<0.05）. Compared with the model group， drug administration relieved the general survival state， the degree of nipple swelling， and the positive behavior in the open field test and reduced the breast thickness （P<0.05）. In addition， drug administration reduced the level of Bcl-2 and increased that of Bax in the breast tissue （P<0.05）， up-regulated the mRNA and protein levels of PTEN， and down-regulated the mRNA and protein levels of PI3K and Akt （P<0.05）. ConclusionGuishao Yunpi decoction can improve the general living conditions and alleviate the mammary gland hyperplasia of rats with the syndrome of liver depression and spleen deficiency， which may be realized by the regulation of the PTEN/PI3K/Akt pathway.

Objective:To observe the clinical efficacy of autologous platelet rich gel (APG) in the treatment of type 2 diabetic foot (DF) patients and the effect of APG on the expression of metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in peripheral blood mononuclear cells (PBMCs).Methods:A total of 62 patients with DF admitted to the Affiliated Hospital of Kangda College of Nanjing Medical University from February 2021 to May 2022 were randomly divided into a control group (30 cases) and an observation group (32 cases) using a random number table method. The control group received ultrasound debridement and dressing change treatment, while the observation group received ultrasound debridement combined with APG treatment. After 6 weeks of treatment, the effective rate, transcutaneous oxygen partial pressure (TcPO 2), and serum tumor necrosis factor- α (TNF-α), interleukin-6 (IL-6), vascular endothelial growth factor (VEGF), hypoxia inducible factor α (HIF-1 α)and the level of MALAT1 expression in PBMCs of the two groups of patients were observed. The Pearson correlation analysis was used to investigate the relationship between the expression change of MALAT (△ MALAT1) and the total effective rate of treatment. Results:The total effective rate of the observation group was higher than that of the control group [93.75%(30/32) vs 73.33%(22/30), P<0.05]. After treatment, the systolic blood pressure (SBP), diastolic blood pressure (DBP), cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), fasting blood glucose (FPG), glycosylated hemoglobin (HbA 1c), urinary microalbumin/creatinine (UACR), uric acid (UA), white blood cells (WBC), TNF- α and IL-6 of both groups had decreased compared to before; HIF-1 α, VEGF and MALAT1 increased compared to before treatment (all P<0.05); After treatment, there was a statistically significant difference in UA, HIF-1α, VEGF, and MALAT1 between the observation group and the control group (all P<0.05). Pearson correlation analysis showed that Δ MALAT1 in DF patients was negatively correlated with TNF -α ( r=-0.61, P=0.02), IL-6 ( r=-0.52, P=0.04), WBC ( r=-0.53, P=0.03), and positively correlated with VEGF ( r=0.58, P=0.03) and HIF-1α ( r=0.54, P=0.03). The total effective rate of DF treatment was higher in the high change group of△ MALAT [88.37%(38/43) vs 73.68%(14/19), P<0.05]. There was no statistically significant difference in the incidence of adverse reactions between the two groups ( P>0.05). Conclusions:APG can significantly upregulate the expression of MALAT, improve wound tissue blood perfusion, wound angiogenesis, and inflammatory response, promote ulcer healing, and changes in MALAT expression can help determine the prognosis of DF.

Balancing the risks and benefits of organophosphate pesticides(OPs)on human and environmental health relies partly on their accurate measurement.A highly sensitive fluorescence anti-quenching multi-residue bio-barcode immunoassay was developed to detect OPs(triazophos,parathion,and chlorpyrifos)in apples,turnips,cabbages,and rice.Gold nanoparticles were functionalized with monoclonal antibodies against the tested OPs.DNA oligonucleotides were complementarily hybridized with an RNA fluorescent label for signal amplification.The detection signals were generated by DNA-RNA hybridization and ribonuclease H dissociation of the fluorophore.The resulting fluorescence signal en-ables multiplexed quantification of triazophos,parathion,and chlorpyrifos residues over the concen-tration range of 0.01-25,0.01-50,and 0.1-50 ng/mL with limits of detection of 0.014,0.011,and 0.126 ng/mL,respectively.The mean recovery ranged between 80.3％and 110.8％with relative standard deviations of 7.3％-17.6％,which correlate well with results obtained by liquid chromatography-tandem mass spectrometry(LC-MS/MS).The proposed bio-barcode immunoassay is stable,reproducible and reliable,and is able to detect low residual levels of multi-residue OPs in agricultural products.