NAC transcription factor ATAF1 negatively modulates the PIF-regulated hypocotyl elongation under a short-day photoperiod.

Day length modulates hypocotyl elongation in seedlings to optimize their overall fitness. Variations in cell growth-associated genes are regulated by several transcription factors. However, the specific transcription factors through which the plant clock increases plant fitness are still being elucidated. In this study, we identified the no apical meristem, Arabidopsis thaliana-activating factor (ATAF-1/2), and cup-shaped cotyledon (NAC) family transcription factor ATAF1 as a novel repressor of hypocotyl elongation under a short-day (SD) photoperiod. Variations in day length profoundly affected the transcriptional and protein levels of ATAF1. ATAF1-deficient mutant exhibited increased hypocotyl length and cell growth-promoting gene expression under SD conditions. Moreover, ATAF1 directly targeted and repressed the expression of the cycling Dof factor 1/5 (CDF1/5), two key transcription factors involved in hypocotyl elongation under SD conditions. Additionally, ATAF1 interacted with and negatively modulated the effects of phytochrome-interacting factor (PIF), thus inhibiting PIF-promoted gene expression and hypocotyl elongation. Taken together, our results revealed ATAF1-PIF as a crucial pair modulating the expression of key transcription factors to facilitate plant growth during day/night cycles under fluctuating light conditions.

An imported malaria case with repeated episodes of neurological syndromes resulting from different Plasmodium species.

BACKGROUND: Imported cerebral malaria (CM) cases in non-endemic areas are often misdiagnosed, which delays treatment. Post-malaria neurological syndrome (PMNS) after recovery from severe malaria can also complicate diagnosis. CASE: We report an imported malaria case from West Africa with two sequential episodes with neurological syndromes within about a month. The first episode was diagnosed as CM with microscopy-positive Plasmodium falciparum infection. The second episode, occurring a month after the recovery from the first CM episode, was consistent with PMNS, since malaria parasites were not detected by microscopy in peripheral blood smears. However, this diagnosis was complicated by the detection of Plasmodium vivax in peripheral blood by PCR, suggesting a potential cause of the second episode by P. vivax. CONCLUSION: This study suggests that PMNS often occurs after severe falciparum malaria. Concurrent P. vivax infection with pathogenic biomass being predominantly extravascular further complicates accurate diagnosis.

Practical NIR Assay Derived from Cyanine to Evaluate Intracellular H2S in Living Cell Imaging.

To monitor the biological function of H2S in real time, this investigation demonstrated the design and synthesis of a novel fluorescent probe integrated with cyanine and 2,4-dinitrophenol for the qualitative and quantitative detection of H2S. An NIR sensitive sensor (FS-HS-1) was provided with a straightforward process. Spectroscopy experiments elucidated that FS-HS-1 could selectively detect H2S in a PBS solution (containing 40% acetonitrile) with a 111-fold fluorescence enhancement at 715 nm (ex. 605 nm). The response towards NaHS occurred in less than 2 min, and the detection limit was confirmed to be as low as 4.47 ± 0.11 nmol/L. Furthermore, the probe is capable of monitoring changes in exogenous H2S concentrations within living cells with confocal and 2P imaging.

Vascular endothelial-derived Von Willebrand factor inhibits lung cancer progression through the αvß3/ERK1/2 axis.

Lung cancer remains a common malignant tumor causing death due to the rapid industrialization and serious pollution of the environment. The Von Willebrand Factor (vWF) protein is an endothelial marker and is widely used to diagnose cancer and other inflammations, however its exact mechanism of action remains largely unexplored. In particular, how it plays two opposing roles in tumor development is not clear. Our study aimed to the impact of endothelial-derived vWF on tumor development by co-culturing human umbilical vein endothelial cells (HUVECs) with lung cancer cells (95D and A549). A knockdown of endothelial-derived vWF assisted lung cancer cell in proliferation, migration and inhibited apoptosis in vitro, while overexpression of endothelial-derived vWF inhibited the proliferation, migration and induced apoptosis of lung cancer cells. The results of further experiments indicated that the vWF secreted by endothelial cells could affect lung cancer cell migration and apoptosis via its binding to integrin αvß3 on the surface of lung cancer cells. Furthermore, a novel finding was the fact that endothelial-derived vWF inhibited lung cancer cell apoptosis by phosphorylating ERK1/2. At the same time, we established experimental lung metastasis model and xenograft model in normal mice and vWF-/- mice, and found that knockout of vWF in mice significantly promoted lung cancer growth and metastasis. In conclusion, our research found that endothelial-derived vWF could directly combine to αvß3 on the exterior of A549 and 95D, thereby mediating lung cancer proliferation, migration and apoptosis and inhibiting the development of lung cancer.

Lung adenocarcinoma-derived vWF promotes tumor metastasis by regulating PHKG1-mediated glycogen metabolism.

Tumor metastasis is a series of complicated biological events. Hematogenous metastasis mediated by von Willebrand factor (vWF) is critical in tumor metastasis. However, the source of vWF and its role in tumor metastasis are controversial, and the further mechanism involved in mediating tumor metastasis is still unclear. In this study, we first demonstrated that lung adenocarcinoma cells could express vWF de novo and promotes tumor metastasis. Through the analysis of transcriptome sequencing, the metastasis promotion effect of vWF may be related to phosphorylase kinase subunit G1 (PHKG1), a catalytic subtype of phosphorylase kinase (PhK). PHKG1 was highly expressed in lung adenocarcinoma patients and led to poor prognosis. Further experiments found that lung adenocarcinoma-derived vWF induced the upregulation of PHKG1 through the PI3K/AKT pathway to promote glycogenolysis. Glycogen was funneled into glycolysis, leading to increased metastasis. Tumor metastasis assayed in vitro and in vivo showed that knockdown of PHKG1 or synergistic injection of phosphorylase inhibition based on the overexpression of vWF could inhibit metastasis. In summary, our research proved that lung adenocarcinoma-derived vWF may mediate tumor metastasis by regulating PHKG1 to promote glycogen metabolism and suggested potential targets for inhibition of lung adenocarcinoma metastasis.

Breast cancer cells-derived Von Willebrand Factor promotes VEGF-A-related angiogenesis through PI3K/Akt-miR-205-5p signaling pathway.

The metastasis and angiogenesis of breast cancer has always been a difficult problem for treatment. It has recently been discovered that Von Willebrand Factor (vWF), in addition to hemostasis, also plays a role in tumor metastasis and angiogenesis. We noticed that besides endothelial cells, breast cancer cells (MDA-MB-231 and MCF-7) could also express vWF. In vitro experiments showed that knocking down vWF inhibited breast cancer cell metastasis. And we found that overexpression of vWF significantly promoted VEGF-A-dependent vascular proliferation in vitro by activating the PI3K/Akt signaling pathway. Further studies indicated that inhibition of PI3K/Akt signaling pathway up-regulated the expression of miR-205-5p, and miR-205-5p could bind to the 3'UTR region of VEGF-A to hinder the production of VEGF-A. Furthermore, when a spontaneous lung metastasis model was established in Balb/c female mice, knockdown of vWF in 4 T1 cells resulted in a decrease in tumor blood vessel density and effectively inhibited lung metastasis, accompanied by a decrease in the expression level of VEGF-A and an increase in the expression level of miR-205-5p. In summary, our findings provide experimental evidence that overexpression of vWF in breast cancer cells down-regulates the expression of miR-205-5p and up-regulates the expression of VEGF-A through the PI3K/Akt signaling pathway, thereby promoting tumor angiogenesis and metastasis.

Semi-Embedding Zn-Co3O4 Derived from Hybrid ZIFs into Wood-Derived Carbon for High-Performance Supercapacitors.

Transition metal oxides (TMOs) can provide high theoretical capacitance due to the change of multiple valence states of transition metals. However, their intrinsic drawbacks, including poor electrical conductivity, lower energy density, and huge volume expansion, will result in the pulverization of electrode materials and restricted electrochemical kinetics, thus leading to poor rate capability and rapid capacity fading. Composite electrodes based on transition metal oxides and carbon-based materials are considered to be promising candidates for overcoming these limitations. Herein, we reported a preparation method of hybrid ZIFs derived Zn-doped Co3O4/carbon (Zn-Co3O4/C-230) particles semi-embedded in wood-derived carbon skeleton for integrated electrodes. A large specific surface area, excellent conductivity, and electrochemical stability provide a larger electrochemical activity and potential window for the electrode. Prepared Zn-Co3O4@CW-230 electrode (0.6 mm thick) displays ultrahigh area specific capacitances of 7.83 and 6.46 F cm-2 at the current densities of 5 and 30 mA cm-2, respectively. Moreover, a symmetric supercapacitor assembled by two identical Zn-Co3O4@CW-230 electrodes delivers a superior area-specific capacitance of 2.61 F cm-2 at the current densities of 5 mA cm-2 and great energy densities of 0.36 mWh cm-2 (6.0 mWh cm-3) at 2.5 mW cm-2, while maintaining 97.3% of initial capacitance over 10,000 cycles. It notably outperforms those of most carbon-based metal oxides, endowing the Zn-Co3O4@CW-230 with extensive prospects for practical application.

A Nanoparticle Delivery of Plasmid Encoding Hepatocyte Growth Factor for Gene Therapy of Silicosis in Mice.

PURPOSE: Silicosis is a serious occupational disease that is characterized by pulmonary infiltrates and fibrosis and is often refractory to current treatments. New therapeutic strategies for silicosis are needed. Hepatocyte growth factor (HGF) is a latent anti-inflammatory and anti-fibrotic growth factor. METHODS: We prepared a polyethyleneimine-polyethylene glycol/pHGF/hyaluronic acid (PEG-PEI/pHGF/HA) nanomaterials loaded with plasmid DNA encoding HGF gene to increase its transfection efficiency. The characterization, including DNA entrapment efficiency, morphology, particle size, and zeta-potential of PEG-PEI/pHGF/HA was studied. And a PEG-PEI/pHGF/HA (N/P=30:1) nanoparticle with low toxicity and high transfection efficiency was used in treatment for silicosis in mice. RESULTS: The results showed that the human HGF expression in the lungs of the mice was increased, and the inflammatory cell infiltration and fibrous collagen deposition was significantly reduced. CONCLUSION: Therefore, PEG-PEI/pHGF/HA nanoparticle warrant further investigation and may be a potential therapeutic strategy for silicosis.

K13-propeller gene polymorphisms of Plasmodium falciparum and the therapeutic effect of artesunate among migrant workers returning to Guangxi, China (2014-2017).

BACKGROUND: The resistance of Plasmodium falciparum to artemisinin has been identified in Asia and some parts of Africa. The drug resistance of P. falciparum will be an obstacle to the successful elimination of malaria by 2025. Whole-genome sequencing of the artemisinin-resistant parasite line revealed mutations on the k13 gene associated with drug resistance in P. falciparum. To understand the artemisinin resistance of the imported P. falciparum cases from Africa, the mutations in the k13 gene in parasites from imported malaria cases in Guangxi Province were detected and the treatment efficiency of artesunate monotherapy was observed. METHODS: DNA was extracted from 319 blood samples from migrant workers with P. falciparum infection who returned to their hometown in Guangxi Province from Africa between 2014 and 2017. The k13-propeller gene was amplified by nested PCR, and sequencing, gene mutation frequency and geographic difference of imported P. falciparum cases were analysed by comparison with the wild-type strain. Of 319 patients, 158 were P. falciparum-infected and were treated with intravenous injection of artesunate and were observed, including the time of asexual stage clearance and the dose of artesunate used. RESULTS: Of the 319 P. falciparum samples, 12 samples had the k13-propeller mutation, and 11 point mutations were detected; 5 were non-synonymous mutations (T474I, A481T, A578S, V603E, G665S) and were not associated with artemisinin resistance. The clinical treatment observation showed that the median (IQR) dose of artesunate for peripheral blood parasite asexual stage clearance was 407.55 (360-510) mg, and the D3 parasite clearance rate was 70.25%, including the five k13-propeller mutations of P. falciparum. After 7 days of treatment, 98.73% of cases were cleared. Two cases were treated with artemisinin for 8 days with a 960-mg dose to completely clear the asexual parasite, but they did not have a mutation in the k13 gene. CONCLUSIONS: Five mutations of the k13-propeller gene in 319 P. falciparum samples from patients returning from Africa were identified. The frequency of the k13-propeller mutants was low, and the mutations were not strongly associated with artemisinin resistance. The median (IQR) dose of artesunate monotherapy in actual clinical treatment to remove asexual parasite stages was 407.55 (360-510) mg, equivalent to D3-D4. Some P. falciparum cases without a k13-propeller mutation showed obvious delayed clearance of the parasite from peripheral blood. Trial registration The diagnosis of malaria and the treatment of malaria-infected patients are the routine work of Centres for Disease Control and Prevention. Information on the patients was conveyed with the patient's approval, and the research aim, methods, risks and benefits of the study were explained in detail to the patients.

Multiple relapses of Plasmodium vivax malaria acquired from West Africa and association with poor metabolizer CYP2D6 variant: a case report.

BACKGROUND: Plasmodium vivax transmission in West Africa, dominant for the Duffy-negative blood group, has been increasingly recognized from both local residents as well as international travelers who contracted P. vivax malaria there. However, the relapsing pattern and sensitivity to antimalarial treatment of P. vivax strains originated from this region are largely unknown. There is evidence that the efficacy of primaquine for radical cure of relapsing malaria depends on host factors such as the hepatic enzyme cytochrome P450 (CYP) 2D6. CASE PRESENTATION: A 49-year-old Chinese man was admitted to the Shanglin County Hospital in Guangxi Province, China, on December 19, 2016, 39 days after he returned from Ghana, where he stayed for one and a half years. He was diagnosed by microscopy as having uncomplicated P. vivax malaria. Treatment included 3 days of intravenous artesunate (420 mg total), and 3 days of chloroquine (1550 mg total), and 8 days of primaquine (180 mg total). Although parasites and symptoms were cleared rapidly and he was malaria-negative for almost two months, he suffered four relapses with relapse intervals ranging from 58 to 232 days. The last relapse occurred at 491 days from his first vivax attack. For the first three relapses, he was treated similarly with chloroquine and primaquine, sometimes supplemented with additional artemisinin combination therapies (ACTs). For the last relapse, he was treated with intravenous artesunate, 3 days of an ACT, and 7 days of azithromycin, and had remained healthy for 330 days. Molecular studies confirmed P. vivax infections for all the episodes. Although this patient was diagnosed to have normal glucose-6-phosphate dehydrogenase (G6PD) activity, his CYP2D6 genotype corresponded to a *2A/*36 allele variant suggesting of an impaired primaquine metabolizer phenotype. CONCLUSIONS: This clinical case suggests that P. vivax malaria originating from West Africa may produce multiple relapses extending beyond one year. The failures of primaquine as an anti-relapse therapy may be attributed to the patient's impaired metabolizer phenotype of the CYP2D6. This highlights the importance of knowing the host G6PD and CYP2D6 activities for effective radical cure of relapsing malaria by primaquine.

Morphological, transcriptional, and metabolic analyses of osmotic-adapted mechanisms of the halophilic Aspergillus montevidensis ZYD4 under hypersaline conditions.

Halophilic fungi in hypersaline habitats require multiple cellular responses for high-salinity adaptation. However, the exact mechanisms behind these adaptation processes remain to be slightly known. The current study is aimed at elucidating the morphological, transcriptomic, and metabolomic changes of the halophilic fungus Aspergillus montevidensis ZYD4 under hypersaline conditions. Under these conditions, the fungus promoted conidia formation and suppressed cleistothecium development. Furthermore, the fungus differentially expressed genes (P < 0.0001) that controlled ion transport, amino acid transport and metabolism, soluble sugar accumulation, fatty acid ß-oxidation, saturated fatty acid synthesis, electron transfer, and oxidative stress tolerance. Additionally, the hypersalinized mycelia widely accumulated metabolites, including amino acids, soluble sugars, saturated fatty acids, and other carbon- and nitrogen-containing compounds. The addition of metabolites-such as neohesperidin, biuret, aspartic acid, alanine, proline, and ornithine-significantly promoted the growth (P ≤ 0.05) and the morphological adaptations of A. montevidensis ZYD4 grown in hypersaline environments. Our study demonstrated that morphological shifts, ion equilibrium, carbon and nitrogen metabolism for solute accumulation, and energy production are vital to halophilic fungi so that they can build tolerance to high-salinity environments.

Chemical constituents from Bletilla striata and their NO production suppression in RAW 264.7 macrophage cells.

A novel glucoside bletilloside A (1) was isolated from the tubers of Bletilla striata, together with seven known compounds (2-8). Their structures were determined on the basis of extensive spectroscopic analyses. All compounds were evaluated for the inhibition on NO production effects in RAW 264.7 macrophage cells, while militarine (4) and dactylorhin A (5) exhibited moderate inhibitory effects.

Gene therapy for neuropathic pain induced by spared nerve injury with naked plasmid encoding hepatocyte growth factor.

BACKGROUND: Neuropathic pain (NP) is a refractory disease in the clinic with a tremendous impact on the quality of life of patients. Gene therapy is a potential strategy for the management of NP. In the present study, we examined the analgesic effect and mechanism of hepatocyte growth factor (HGF) in vitro and in vivo. METHODS: We examined the proinflammatroy gene changes in lipopolysaccharide (LPS)-induced microglia BV2 cells with a quantitative real-time polymerase chain reaction of interleukin (IL)-1ß, IL-6, tumor necrosis factor (TNF)-α and inducible nitric oxide synthase (iNOS). Mechanical stimulation tests were performed five times at 5-min intervals to assess pain thresholds using Von Frey Hair in mice following spared nerve injury (SNI). The glial cell activation of spinal cord was examined by western blotting. Statistical significance was determined by a Tukey's test and a paired t-test. RESULTS: We found that recombinant human HGF protein suppressed LPS-induced BV2 cell activation in vitro, marked by the down-regulation of IL-1ß, IL-6, TNF-α and iNOS expression, as well as decrease of nitric oxide production. Moreover, intrathecal injection of naked plasmid encoding HGF gene (pUDK-HGF) significantly attenuated SNI-induced pain behaviors in mice by direct inhibition of spinal cord microglia and astrocyte activation. CONCLUSIONS: The results of the present study indicate that pUDK-HGF can reduce cytotoxicity products released from activated glial cells, which may provide a promising therapeutic strategy for treating NP.

Gram-scale production of plasmid pUDK-HGF with current good manufacturing practices for gene therapy of critical limb ischemia.

The demand of a plasmid encoding human hepatocyte growth factor gene (pUDK-HGF) in large quantities at high purity and concentration has increased for gene therapy of critical limb ischemia (CLI) in clinical trials. In this article, we produced pUDK-HGF in compliance with current good manufacturing practices at gram scale. The process included a 50-L batch fermentation, continuous alkaline lysis, and integrated three-step chromatography on Sepharose 6 Fast Flow, PlasmidSelect Xtra, and Source 15Q. The production process has been scaled up to yield 4.24 ± 0.41 g of pharmaceutical pUDK-HGF from 1.0 kg bacterial cell paste and the overall yield reached range from 58.37 to 66.70%. The final pUDK-HGF product exhibited high purity with supercoiled percentage of > 95.8% and undetectable residual RNA, contaminated protein, and bacterial endotoxin. The phase I clinical study indicates that intramuscular injection of pUDK-HGF is safe, well tolerated, and may provide symptomatic relief to CLI patients. These results show that our manufacturing process of pUDK-HGF is efficient in producing pharmaceutical-grade plasmid DNA and is safe for clinical applications.

[Enhancing the glycerol utilization of engineered yeast increases its bisabolene production].

Bisabolene is a compound commonly found in essential oils of various plants. It has a broad application in sectors such as chemical, pharmaceutical, and health-care products. This study focuses on modifying the glycerol metabolism pathway to obtain a high bisabolene-producing strain of Saccharomyces cerevisiae. To achieve this, the glycerol transporter gene PtFPS2 from Pachysolen tannophilus and the glycerol dehydrogenase gene Opgdh from Ogataea parapolymorpha were overexpressed in engineered yeast YS036, which was equipped with a GAL promoters-enhanced mevalonic acid pathway. Additionally, the glucose-inhibiting transcription factor MIG1 was knocked out to reduce glucose inhibition. The results showed that the GAL promoter transcription levels of the recombinant yeast strains increased, and the co-utilization of sucrose and glycerol was further improved in MIG1-knockout strain. Moreover, the maximum yield of bisabolene in shaking flask fermentation increased to 866.7 mg/L, an 82.2% increase compared to that of the original strain. By modifying the metabolic pathway of carbon sources, the yield of bisabolene was considerably improved. This study offers an effective strategy for enhancing the yield of terpene compounds in engineered yeast.

[Empagliflozin inhibits inflammatory response and alleviates renal injury in type 2 diabetic rats by up-regulating the expression of exchange protein 1 directly activated by cAMP (Epac1)].

Objective To observe the therapeutic effect of empagliflozin (EM) on renal injury in rats with type 2 diabetes mellitus (T2DM), and to explore its possible mechanism. Methods Male SD rats were randomly divided into a normal control (NC) group, a T2DM group, and an EM group, with 6 rats in each group. T2DM models were established by an intraperitoneal injection of streptozotocin (STZ) in the T2DM and EM groups. Fasting blood glucose (FBG) levels and body mass of rats in each group were recorded. The EM group received EM solution through intragastric administration, while the other two groups were given an equivalent volume of sodium carboxymethyl cellulose solution through intragastric administration for 12 weeks. After the body mass and FBG levels were recorded, the rats were sacrificed and blood samples from the abdominal aorta and kidney tissues were collected. Serum creatinine (Scr), blood urea nitrogen (BUN), uric acid (UA), triglyceride (TG) and total cholesterol (TC) were detected by automatic biochemical analyzer. Masson, PAS and HE staining were used to assess histological changes in the kidneys, and a transmission electron microscopy was used to observe ultrastructural changes. Immunohistochemical staining was used to detect the expression and distribution of exchange protein 1 directly activated by cAMP(Epac1), TNF-α, IL-1ß, and IL-18 in renal tissue of rats. Results Compared with the NC group, the rats in T2DM group showed a decrease in body mass, a significant increase in the levels of FBG, Scr, BUN, UA, TC, and TG, thickened glomerular basement membrane, foot process fusion of podocytes, disordered cell arrangement and loss of endothelial cell fenestrations. The expression level of Epac1 decreased, while the expression levels of TNF-α, IL-1ß, and IL-18 significantly increased. Compared with the T2DM group, the rats in the EM group showed an increase in body mass, significantly decreased levels of FBG, Scr, BUN, UA, TC, and TG, reduced renal injury, increased expression level of Epac1, and significantly decreased expression levels of TNF-α, IL-1ß, and IL-18. Conclusion EM can improve renal injury in T2DM rats by up-regulating Epac1 expression to inhibit inflammatory response.

Osmanthus fragrans Flavonoid Extract Inhibits Adipogenesis and Induces Beiging in 3T3-L1 Adipocytes.

Osmanthus fragrans has a long history of cultivation in Asia and is widely used in food production for its unique aroma, which has important cultural and economic values. It is rich in flavonoids with diverse pharmacological properties, such as antioxidant, anti-tumor, and anti-lipid activities. However, little is known regarding the effects of Osmanthus fragrans flavonoid extract (OFFE) on adipogenesis and pre-adipocyte transdifferentiation. Herein, this research aimed to investigate the effect of OFFE on the differentiation, adipogenesis, and beiging of 3T3-L1 adipocytes and to elucidate the underlying mechanism. Results showed that OFFE inhibited adipogenesis, reduced intracellular reactive oxygen species levels in mature adipocytes, and promoted mitochondrial biogenesis as well as beiging/browning in 3T3-L1 adipocytes. This effect was accompanied by increased mRNA and protein levels of the brown adipose-specific marker gene Pgc-1a, and the upregulation of the expression of UCP1, Cox7A1, and Cox8B. Moreover, the research observed a dose-dependent reduction in the mRNA expression of adipogenic genes (C/EBPα, GLUT-4, SREBP-1C, and FASN) with increasing concentrations of OFFE. Additionally, OFFE activated the AMPK signaling pathway to inhibit adipogenesis. These findings elucidate that OFFE has an inhibitory effect on adipogenesis and promotes browning in 3T3-L1 adipocytes, which lays the foundation for further investigation of the lipid-lowering mechanism of OFFE in vivo in the future.

Local Multiple-site Injections of a Plasmid Encoding Human MnSOD Mitigate Radiation-induced Skin Injury by Inhibiting Ferroptosis.

BACKGROUND: Most patients who undergo radiotherapy develop radiation skin injury, for which effective treatment is urgently needed. MnSOD defends against reactive oxygen species (ROS) damage and may be valuable for treating radiation-induced injury. Here, we (i) investigated the therapeutic and preventive effects of local multiple-site injections of a plasmid, encoding human MnSOD, on radiation-induced skin injury in rats and (ii) explored the mechanism underlying the protective effects of pMnSOD. METHODS: The recombinant plasmid (pMnSOD) was constructed with human cytomegalovirus (CMV) promoter and pUC-ori. The protective effects of pMnSOD against 20-Gy X-ray irradiation were evaluated in human keratinocytes (HaCaT cells) by determining cell viability, ROS levels, and ferroptosisrelated gene expression. In therapeutic treatment, rats received local multiple-site injections of pMnSOD on days 12, 19, and 21 after 40-Gy γ-ray irradiation. In preventive treatment, rats received pMnSOD injections on day -3 pre-irradiation and on day 4 post-irradiation. The skin injuries were evaluated based on the injury score and pathological examination, and ferroptosis-related gene expression was determined. RESULTS: In irradiated HaCaT cells, pMnSOD transfection resulted in an increased SOD2 expression, reduced intracellular ROS levels, and increased cell viability. Moreover, GPX4 and SLC7A11 expression was significantly upregulated, and erastin-induced ferroptosis was inhibited in HaCaT cells. In the therapeutic and prevention treatment experiments, pMnSOD administration produced local SOD protein expression and evidently promoted the healing of radiation-induced skin injury. In the therapeutic treatment experiments, the injury score in the high-dose pMnSOD group was significantly lower than in the PBS group on day 33 post-irradiation (1.50 vs. 2.80, P < 0.05). In the prevention treatment experiments, the skin injury scores were much lower in the pMnSOD administration groups than in the PBS group from day 21 to day 34. GPX4, SLC7A11, and Bcl-2 were upregulated in irradiated skin tissues after pMnSOD treatment, while ACSL4 was downregulated. CONCLUSION: The present study provides evidence that the protective effects of MnSOD in irradiated HaCaT cells may be related to the inhibition of ferroptosis. The multi-site injections of pMnSOD had clear therapeutic and preventive effects on radiation-induced skin injury in rats. pMnSOD may have therapeutic value for the treatment of radiation-induced skin injury.

A recombinant plasmid encoding human hepatocyte growth factor promotes healing of combined radiation-trauma skin injury involved in regulating Nrf2 pathway in mice.

Combined radiation-trauma skin injury represents a severe and intractable condition that urgently requires effective therapeutic interventions. In this context, hepatocyte growth factor (HGF), a multifunctional growth factor with regulating cell survival, angiogenesis, anti-inflammation and antioxidation, may be valuable for the treatment of combined radiation-trauma injury. This study investigated the protective effects of a recombinant plasmid encoding human HGF (pHGF) on irradiated human immortalized keratinocytes (HaCaT) cells in vitro, and its capability to promote the healing of combined radiation-trauma injuries in mice. The pHGF radioprotection on irradiated HaCaT cells in vitro was assessed by cell viability, the expression of Nrf2, Bcl-2 and Bax, as well as the secretion of inflammatory cytokines. In vivo therapeutic treatment, the irradiated mice with full-thickness skin wounds received pHGF local injection. The injuries were appraised based on relative wound area, pathology, immunohistochemical detection, terminal deoxynucleotidyl transferase dUTP nick end labelling assay and cytokine content. The transfection of pHGF increased the cell viability and Nrf2 expression in irradiated HaCaT cells. pHGF also significantly upregulated Bcl-2 expression, decreased the Bax/Bcl-2 ratio and inhibited the expression of interleukin-1ß and tumor necrosis factor-α in irradiated cells. Local pHGF injection in vivo caused high HGF protein expression and noticeable accelerated healing of combined radiation-trauma injury. Moreover, pHGF administration upregulated Nrf2, vascular endothelial growth factor, Bcl-2 expression, downregulated Bax expression and mitigated inflammatory response. In conclusion, the protective effect of pHGF may be related to inhibiting apoptosis and inflammation involving by upregulating Nrf2. Local pHGF injection distinctly promoted the healing of combined radiation-trauma injury and demonstrates potential as a gene therapy intervention for combined radiation-trauma injury in clinic.

Analysis of alterations in serum vitamins and correlations with gut microbiome, microbial metabolomics in patients with sepsis.

BACKGROUND: Vitamins are essential micronutrients that play key roles in many biological pathways associated with sepsis. The gut microbiome plays a pivotal role in the progression of sepsis and may contribute to the onset of multi-organ dysfunction syndrome (MODS). The aim of this study was to investigate the changes in serum vitamins, and their correlation with intestinal flora and metabolomic profiles in patients with sepsis. METHODS: The serum levels of vitamins were determined by Ultra Performance Liquid Chromatography (UPLC). 16S rRNA gene sequencing and Liquid Chromatography-tandem Mass Spectrometry (LC-MS/MS) targeted metabolomics were used for microbiome and metabolome analysis. RESULTS: In the training cohort: After univariate, multivariate (OPLS-DA) and Spearman analyses, it was concluded that vitamin levels of 25 (OH) VD3 and (VD2 + VD3), as well as vitamins A and B9, differed significantly among healthy controls (HC), non-septic critical patients (NS), and sepsis patients (SS) (P < 0.05). The validation cohort confirmed the differential vitamin findings from the training cohort. Moreover, analyses of gut flora and metabolites in septic patients and healthy individuals revealed differential flora, metabolites, and metabolic pathways that were linked to alterations in serum vitamin levels. We found for the first time that vitamin B9 was negatively correlated with g_Sellimonas. CONCLUSION: Sepsis patients exhibited significantly lower levels of 25 (OH) VD3 and (VD2 + VD3), vitamins A and B9, which hold potential as predictive markers for sepsis prognosis. The changes in these vitamins may be associated with inflammatory factors, oxidative stress, and changes in gut flora.