Objective To explore the mechanism by which zinc-regulated transporters, iron-regulated transporter-likeprotein 4 (ZIP4) regulates glycolysis and its impact on tumor progression in cholangiocarcinoma (CCA), providing a theoretical basis for targeted therapy of CCA. Methods ZIP4 expression in CCA was analyzed using the GEPIA database. Immuno-histochemistry (IHC) was used to detect ZIP4 expression in 20 paired CCA and adjacent non-tumor tissues. Stable ZIP4-overexpressing CCA cell lines (ZIP4-OE) were established. Gene set enrichment analysis was used to screen differentially expressed genes and pathways in ZIP-OE CCA cells. ZIP4, N-myc proto-oncogene protein (MYCN), and histone-lysine N-methyltransferase 2E (KMT2E) were knocked down using small interfering RNAs (siRNAs). The expression of glycolysis-related gene (glucose transporter 1 [Glut1], hexokinase 2 [HK2], and lactate dehydrogenase A [LDHA]) was measured by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). Glycolytic activity was assessed by measuring the extracellular acidification rate (ECAR). Cell proliferation was evaluated using colony formation assays, and cell migration was assessed using Transwell assays. A xenograft mouse model was constructed to examine CCA tumor growth. Protein levels of ZIP4, KMT2E, H3K4me3 (tri-methylation of lysine 4 on histone H3), and MYCN were detected by Western blotting. Results GEPIA database analysis and IHC results confirmed significantly higher ZIP4 expression levels in CCA tissues compared to adjacent non-tumor tissues (P＜0.05). Compared to the control group, the ZIP4-OE group exhibited a significantly increased ECAR, along with significantly enhanced proliferation and migration abilities (P＜0.01). Conversely, knockdown of ZIP4 suppressed CCA cells proliferation and migration. GEPIA analysis indicated that ZIP4 upregulates the transcription of oncogene MYCN, as well as glycolysis-related genes. Knockdown of MYCN abolished the ZIP4 overexpression-induced upregulation of Glut1, HK2, and LDHA gene transcription, reduced glycolysis, and significantly inhibited CCA cell proliferation and migration (P＜0.05). Mechanistic studies demonstrated that ZIP4 increases H3K4me3 level via KMT2E, leading to MYCN transcription. Knockdown of KMT2E in CCA cells suppressed the ZIP4 overexpression-induced enhancement in H3K4me3 modification, resulting in MYCN downregulation and significantly reduced CCA cells proliferation and migration (P＜0.05). Conclusions ZIP4 upregulates H3K4me3 modification through KMT2E, which recruits transcription factors to activate the transcription of MYCN. This subsequently enhances cellular glycolysis and promotes the proliferation and migration of CCA cells.

Hypoxic-ischemic (HI) brain damage poses a high risk of death or lifelong disability, yet effective treatments remain elusive. Here, we demonstrated that miR-100-5p levels in the lesioned cortex increased after HI insult in neonatal mice. Knockdown of miR-100-5p expression in the brain attenuated brain injury and promoted functional recovery, through inhibiting the cleaved-caspase-3 level, microglia activation, and the release of proinflammation cytokines following HI injury. Engineered extracellular vesicles (EVs) containing neuron-targeting rabies virus glycoprotein (RVG) and miR-100-5p antagonists (RVG-EVs-Antagomir) selectively targeted brain lesions and reduced miR-100-5p levels after intranasal delivery. Both pre- and post-HI administration showed therapeutic benefits. Mechanistically, we identified protein phosphatase 3 catalytic subunit alpha (Ppp3ca) as a novel candidate target gene of miR-100-5p, inhibiting c-Fos expression and neuronal apoptosis following HI insult. In conclusion, our non-invasive method using engineered EVs to deliver miR-100-5p antagomirs to the brain significantly improves functional recovery after HI injury by targeting Ppp3ca to suppress neuronal apoptosis.
Animals ; MicroRNAs/metabolism* ; Extracellular Vesicles/metabolism* ; Mice ; Recovery of Function/physiology* ; Hypoxia-Ischemia, Brain/therapy* ; Mice, Inbred C57BL ; Antagomirs/administration & dosage* ; Male ; Animals, Newborn ; Apoptosis/drug effects* ; Brain Injuries/metabolism* ; Glycoproteins ; Peptide Fragments ; Viral Proteins

Objective:To explore the causal relationship between thyroid dysfunction and sepsis based on the bidirectional two-sample Mendelian randomization (MR) method.Methods:The genome-wide association study (GWAS) dataset were selected to screen single nucleotide polymorphisms (SNP) associated with thyroid dysfunction as instrumental variable (IV) for genetic variation, using hypothyroidism and hyperthyroidism as exposure factor and sepsis as outcome factor. Potential causal relationship between thyroid dysfunction and sepsis was analyzed using a bidirectional two-sample MR method primary analysis method of inverse-variance weighted (IVW). Potential pleiotropic analysis of SNP was performed using the MR Egger regression intercept test. Sensitivity analysis was performed using the "leave one out" test. Reverse MR method was used to prove the causal relationship.Results:The GWAS data were screened based on the three main assumptions of MR, resulting in 101 SNP strongly associated with hypothyroidism and 10 SNP strongly associated with hyperthyroidism entering the MR analysis. The results of the MR using the IVW method showed that the risk of sepsis in individuals with hypothyroidism was 2.293 times higher than those without hypothyroidism [odds ratio ( OR) = 2.293, 95% confidence interval (95% CI) was 1.199-4.382, P = 0.012]. There was no significant difference in the risk of sepsis between hyperthyroid and non-hyperthyroid populations ( OR = 1.049, 95% CI was 0.999-1.100, P = 0.560). MR Egger regression intercept test showed that the included SNP did not have pleiotropy, and the MR-PRESSO test did not find outliers. Sensitivity analysis suggested that the results of MR were stable. The results of the reverse MR analysis showed that the reverse causal relationship between hyperthyroidism and sepsis was not proved ( OR = 0.996, 95% CI was 0.988-1.004, P = 0.338), which further confirmed the robust MR analysis result. Conclusion:The results of the bidirectional two-sample MR analysis show that hypothyroidism can increase the risk of sepsis onset, while there is no causal relationship between hyperthyroidism and sepsis.

Proteins in the human body are usually made of 20 natural amino acids.Through different amino acid combinations and isomerization, proteins of diverse functions are built.An emerging genetic code expansion technology can introduce unnatural amino acids into specific sites of target protein, endowing the protein with new biological characteristics including covalently binding with proximal proteins, carrying fluorescence, and mimicking specific protein post-translational modifications.In this paper, based on the structure and function of unnatural amino acids, the applications of different types of unnatural amino acids in regulating protein''s stability, studying protein''s conformation, expression level, and localization, and uncovering heretofore unknown protein-protein interactions were reviewed.Besides, genetic code expansion of unnatural amino acids is anticipated to find broad utilities in biomedicine by bringing new ideas and methods to the design and optimization of biologics.

Rheumatoid arthritis (RA) is an autoimmune disease and is mainly characterized by abnormal proliferation of fibroblast-like synoviocytes (FLS). The up-regulated cellular membrane expression of G protein coupled receptor kinase 2 (GRK2) of FLS plays a critical role in RA progression, the increase of GRK2 translocation activity promotes dysfunctional prostaglandin E4 receptor (EP4) signaling and FLS abnormal proliferation. Recently, although our group found that paeoniflorin-6'-

Objective:To investigate the short-term outcomes of laparoscopic simultaneous resection of primary colorectal cancer and liver metastases in patients with resectable synchronous colorectal liver metastases (SCRLM).Methods:A descriptive case series study was performed. Clinicopathological data of patients with SCRLM who underwent laparoscopic simultaneous resection of colorectal cancer and liver metastases in Zhongshan Hospital between December 2015 and September 2018 were retrieved from a prospective colorectal cancer database. Perioperative presentations and short-term outcomes were analyzed.Results:A total of 53 patients were enrolled with average age of(61.7±11.3) years. Among them, 32 were male (60.4%) and 21 were female (39.6%). Twenty-five patients (47.2%) were American Society of Anesthesiologists (ASA) grade I and 28 (52.8%) were grade II. All the patients completed laparoscopic simultaneous resection without conversion. The average operation time was (320.2±114.5) min. The estimated blood loss was 150.0 (45.0-2000.0) ml, and only 2 cases (3.8%) received intraoperative transfusion. Postoperative pathologic results revealed that the average primary tumor size was (5.4±1.9) cm; 4 cases (7.5%) were T1-2 stage and 48 cases (90.6%) were T3-4 stage; 40 patients (75.5%) had lymph node metastasis; 19 (35.8%) had vascular involvement; 24 (45.3%) had neural invasion. The median number of liver metastases was 1.0 (1-8), and the average size of largest liver metastases was (3.0±1.9) cm. The median margin of liver metastases was 1.0 (0.1-3.5) cm, and only 1 case was R1 resection. The average time to the first postoperative flatus was (67.9±28.9) h, and the average time to the liquid diet was (107.0±33.8) h. The average postoperative indwelling catheterization time was (85.6±56.4) h. The average postoperative hospital stay was (9.2±4.4) d, and the average cost was (82±26) thousand RMB. No death within postoperative 30-day was found. The morbidity of postoperative complication was 32.1% (17/53) and 3 patients developed grade III to IV complications which were improved by conservative treatment. The median follow-up period was 23.2 months. During follow-up, 19 patients (35.8%) developed recurrence or metastasis, and 4 (7.5%) died. The 1- and 2-year disease-free survival (DFS) rates were 68% and 47% respectively, and the 1- and 2-year overall survival rates were 95% and 86% respectively.Conclusions:Laparoscopic simultaneous resection of primary colorectal cancer and liver metastases is safe and feasible in selected patients with SCRLM. Postoperative intestinal function recovery is enhanced, and morbidity and oncological outcomes are acceptable.

Objective:To evaluate the safety and efficacy of eltrombopag combined with immunosuppressive therapy in patients with aplastic anemia (AA) in China.Methods:We investigated and analyzed the clinical data of AA patients from 14 hematological treatment centers who were treated with oral eltrombopag for at least 3 mon.Results:We enrolled 56 AA patients, including 19 treatment-na?ve patients and 37 IST-refractory patients. The median administration period for eltrombopag was 7 (3-31) months, and the median maximum stable dosage was 75 mg/d (50-150 mg/d) . The 3-month hematological response (HR) rate was 60%, and the complete response (CR) rate was 30% in 10 SAA patients who were treated with first-line eltrombopag and standard IST (ATG+CsA) . Eight of 9 eltrombopag and CsA ± androgen first-line treated SAA patients responded (8/9, 89%) and 4 (44%) gave CR. The overall HR and CR rates were 79% and 52.6%, respectively, among these 19 patients by the end of the follow-up period. Of the 19 AA patients who were refractory to CsA ± androgen, 11 achieved HR (57.9%) at 3 mon, and the best HR rate was 44% in standard IST (ATG+CsA) refractory 18 patients after eltrombopag treatment. Fifty-one percent of the patients experienced mild or moderate adverse events, and gastrointestinal discomfort was the most common adverse effect reported by the study subjects.Conclusion:Adding Eltrombopag in first-line IST can accelerate the acquisition and improve the quality of hematological responses in AA patients. AA with relatively more residual hematopoietic cells may be well treated with eltrombopag and non-ATG IST. Eltrombopag can be used as salvage therapy for CsA±androgen refractory patients. Eltrombopag was generally safe and well tolerated by AA patients in China.

Objective:To investigate the short-term outcomes of laparoscopic simultaneous resection of primary colorectal cancer and liver metastases in patients with resectable synchronous colorectal liver metastases (SCRLM).Methods:A descriptive case series study was performed. Clinicopathological data of patients with SCRLM who underwent laparoscopic simultaneous resection of colorectal cancer and liver metastases in Zhongshan Hospital between December 2015 and September 2018 were retrieved from a prospective colorectal cancer database. Perioperative presentations and short-term outcomes were analyzed.Results:A total of 53 patients were enrolled with average age of(61.7±11.3) years. Among them, 32 were male (60.4%) and 21 were female (39.6%). Twenty-five patients (47.2%) were American Society of Anesthesiologists (ASA) grade I and 28 (52.8%) were grade II. All the patients completed laparoscopic simultaneous resection without conversion. The average operation time was (320.2±114.5) min. The estimated blood loss was 150.0 (45.0-2000.0) ml, and only 2 cases (3.8%) received intraoperative transfusion. Postoperative pathologic results revealed that the average primary tumor size was (5.4±1.9) cm; 4 cases (7.5%) were T1-2 stage and 48 cases (90.6%) were T3-4 stage; 40 patients (75.5%) had lymph node metastasis; 19 (35.8%) had vascular involvement; 24 (45.3%) had neural invasion. The median number of liver metastases was 1.0 (1-8), and the average size of largest liver metastases was (3.0±1.9) cm. The median margin of liver metastases was 1.0 (0.1-3.5) cm, and only 1 case was R1 resection. The average time to the first postoperative flatus was (67.9±28.9) h, and the average time to the liquid diet was (107.0±33.8) h. The average postoperative indwelling catheterization time was (85.6±56.4) h. The average postoperative hospital stay was (9.2±4.4) d, and the average cost was (82±26) thousand RMB. No death within postoperative 30-day was found. The morbidity of postoperative complication was 32.1% (17/53) and 3 patients developed grade III to IV complications which were improved by conservative treatment. The median follow-up period was 23.2 months. During follow-up, 19 patients (35.8%) developed recurrence or metastasis, and 4 (7.5%) died. The 1- and 2-year disease-free survival (DFS) rates were 68% and 47% respectively, and the 1- and 2-year overall survival rates were 95% and 86% respectively.Conclusions:Laparoscopic simultaneous resection of primary colorectal cancer and liver metastases is safe and feasible in selected patients with SCRLM. Postoperative intestinal function recovery is enhanced, and morbidity and oncological outcomes are acceptable.

Objective The aim of this study was to investigate the expressions of and clinical significance of F-box/WD-40 domain protein 10(FBXW10) as well as the expression of cell cycle protein cyclin E( cyclin E) in renal clear cell carcinoma. Methods Immunohistochemistry SP method was used to detect the expressions of FBXW10 and cyclin E protein in 60 cases of renal clear cell carcinoma and 20 cases of adjacent normal tissues. The relationship between the expressions of FBXW10 and cyclin E,and the clinical pathological characteristics was analyzed. Results The expression rates of FBXW10 and cyclin E protein in renal clear cell carcinoma were 40. 0% ,70. 0% ,respectively and adjacent normal tissues were 55. 0% and 25. 0% ( P<0. 05). The expression of FBXW10 was correlated with the histologic grade of renal clear cell carcinoma(P=0. 041),histologic grade( P=0. 030);the ex-pression of cyclin E was correlated with the pathological tumor stage of clear cell renal cell carcinoma(P=0. 005),degree of differen-tiation(P=0. 035),and distant metastasis(P=0. 011). There was a significant correlation between the expressions of FBXW10 and cyclin E in renal clear cell carcinoma(r=0. 533,P<0. 001). Conclusion FBXW10 and cyclin E may play important roles in the development of renal clear cell carcinoma.

Objective: To evaluate the immunogenicity of split influenza H1N1 vaccine formulated with an oil-in-water nano-emulsion adjuvant in aged mice and young mice. Methods: A nano-emulsion adjuvant formulated split influenza H1N1 vaccine was used to immunize aged and young mice through intramuscular injection. Each mouse was immunized with 0.012 μg of hemagglutinin (HA) twice with an interval of 28 d. Hemagglutination inhibition (HI) titers in serum were measured 27 d after first immunization. Serum HI, IgG1 and IgG2a titers were detected 14 d after the last immunization. No adjuvant-formulated vaccine and normal saline (NS) were used to set up control groups. Virus challenge test was carried out using 10 times the median lethal dose (LD₅₀) of A/Puerto Rico/8/34 (H1N1) strain two weeks after the last immunization and the protective effects were assessed through measuring the dynamic changes in body weight and survival rate. Results: Higher levels of serum HI, IgG1 and IgG2a antibodies and higher HI antibody conversion rates were induced in the adjuvant groups, especially in the aged mice group, than in the control groups. Nano-emulsion adjuvant improved the immunogenicity of HA and mouse immunity to A/Puerto Rico/8/34 (H1N1). Conclusions Nano-emulsion adjuvant could enhance the immunogenicity of influenza antigens, especially in aged mice.