A DNA/Upconversion Nanoparticle Complex Enables Controlled Co-Delivery of CRISPR-Cas9 and Photodynamic Agents for Synergistic Cancer Therapy.

Photodynamic therapy (PDT) depends on the light-irradiated exciting of photosensitizer (PS) to generate reactive oxygen species (ROS), which faces challenges and limitations in hypoxia and antioxidant response of cancer cells, and limited tissue-penetration of light. Herein, a multifunctional DNA/upconversion nanoparticles (UCNPs) complex is developed which enables controlled co-delivery of CRISPR-Cas9, hemin, and protoporphyrin (PP) for synergistic PDT. An ultralong single-stranded DNA (ssDNA) is prepared via rolling circle amplification (RCA), which contains recognition sequences of single guide RNA (sgRNA) for loading Cas9 ribonucleoprotein (RNP), G-quadruplex sequences for loading hemin and PP, and linker sequences for combining UCNP. Cas9 RNP cleaves the antioxidant regulator nuclear factor E2-related factor 2 (Nrf2), improving the sensitivity of cancer cells to ROS, and enhancing the synergistic PDT effect. The G-quadruplex/hemin DNAzyme mimicks horseradish peroxidase (HRP) to catalyze the endogenous H2O2 to O2, overcoming hypoxia condition in tumors. The introduced UCNP converts NIR irradiation with deep tissue penetration to light with shorter wavelength, exciting PP to transform the abundant O2 to 1O2. The integration of gene editing and PDT allows substantial accumulation of 1O2 in cancer cells for enhanced cell apoptosis, and this synergistic PDT has shown remarkable therapeutic efficacy in a breast cancer mouse model.

Progress of nanopreparation technology applied to volatile oil drug delivery systems.

Traditional Chinese medicine volatile oil has a long history and possesses extensive pharmacological activity. However, volatile oils have characteristics such as strong volatility, poor water solubility, low bioavailability, and poor targeting, which limit their application. The use of volatile oil nano drug delivery systems can effectively improve the drawbacks of volatile oils, enhance their bioavailability and chemical stability, and reduce their volatility and toxicity. This article first introduces the limitations of the components of traditional Chinese medicine volatile oils, discusses the main classifications and latest developments of volatile oil nano formulations, and briefly describes the preparation methods of traditional Chinese medicine volatile oil nano formulations. Secondly, the limitations of nano formulation technology are discussed, along with future challenges and prospects. A deeper understanding of the role of nanotechnology in traditional Chinese medicine volatile oils will contribute to the modernization of volatile oils and broaden their application value.

Network pharmacology and molecular docking of endogenous active metabolites in diabetic kidney disease.

OBJECTIVES: Network pharmacology and molecular docking were used to predict endogenous active metabolites with protective effects in diabetic kidney disease (DKD). METHODS: We utilized metabolomics to screen differentially expressed metabolites in kidney tissues of mice with type 2 DKD and predicted potential targets using relevant databases. The interaction network between endogenous active metabolites and target proteins was established by integrating differentially expressed metabolites and proteins associated with DKD identified through proteomics. Gene ontology (GO) and signaling pathway enrichment analysis were performed. The biological functions of the active candidate metabolites and their effects on downstream pathways were also verified. RESULTS: Metabolomics revealed 130 differentially expressed metabolites. Through co-expression network analysis coupled with the investigation of differentially expressed proteins in proteomics, 2-hydroxyphenylpropionylglycine (2-HPG) emerged as a key regulator of DKD. 2-HPG was found to modulate the progression of DKD by regulating the conformation and activity of synaptophysin 1 (SYNJ1), with a correlation coefficient of 0.974. In vivo experiments revealed that SYNJ1 expression was significantly downregulated in the Macroalbuminuria Group compared to the Control Group and negatively correlated with proteinuria (r = -0.7137), indicating its important role in DKD progression. Immunofluorescence demonstrated that treatment with 2-HPG restores the expression of the foot process marker protein Wilms tumor-1 (WT-1) in podocytes injured by high glucose levels. Western blot and polymerase chain reaction support the involvement of SYNJ1 in this process. CONCLUSIONS: This study demonstrated the significance of the 2-HPG/SYNJ1 signaling axis in safeguarding the foot process of podocytes in DKD.

Framework-Hotspot Enhanced Trans Cleavage of CRISPR-Cas12a for Clinical Samples Detection.

The trans-cleavage property of CRISPR-Cas12a system makes it an excellent tool for disease diagnosis. Nevertheless, most methods based on CRISPR-Cas system still require pre-amplification of the target to achieve the desired detection sensitivity. Here we generate Framework-Hotspot reporters (FHRs) with different local densities to investigate their effect on trans-cleavage activity of Cas12a. We find that the cleavage efficiency increases and the cleavage rate accelerates with increasing reporter density. We further construct a modular sensing platform with CRISPR-Cas12a-based target recognition and FHR-based signal transduction. Encouragingly, this modular platform enables sensitive (100 fM) and rapid (<15 min) detection of pathogen nucleic acids without pre-amplification, as well as detection of tumor protein markers in clinical samples. The design provides a facile strategy for enhanced trans cleavage of Cas12a, which accelerates and broadens its applications in biosensing.

The Function of Spag6 to Repair Brain Edema Damage After Cerebral Ischemic Stroke-reperfusion.

Sperm associated antigen 6 (Spag6) is the PF16 homolog of Chlamydomonas and participates in the regulation of cilia movement. Studies have shown that Spag6 is expressed in the brain, and its loss will lead to cerebral edema caused by a defect in motor cilium function in ependymal cells. However, it has not been reported whether the limited or extensive cerebral edema resulting from ischemic strokes is related to the expression regulation of Spag6. Therefore, this study aimed to investigate the effect and related mechanism of Spag6 in alleviating Cerebral Ischemic stroke-reperfusion (CIS/R) damage. Our experimental results showed that Spag6 overexpression alleviated CIS/R-mediated motor cilia structural disorder, improved cerebral edema, inhibited nerve injuries in rats with cerebral ischemia, and alleviated synaptic and dendritic spinal injuries by regulating the expressions of synaptic-related proteins such as CaMKII, PSD95, and CREB. Based on significant changes in PI3K/AKT-mTOR signaling pathway activity after CIS/R determination, we determined that Spag6 regulates the abnormal expression of CIS/R-induced inflammatory factors NF-κB, NLRP3, IL-10, and the autophagy-related proteins Beclin-1, LC3, and P62 by activating the PI3K/AKT-mTOR signaling pathway. This inhibits inflammation and autophagy in the brain tissue. In summary, this study revealed that Spag6 alleviates brain edema damage after CIS/R by maintaining the structural function of the motor cilium, regulating the PI3K/AKT-mTOR signaling pathway, and inhibiting inflammation and autophagy reaction.

Investigating the effect of dehydromiltirone on septic AKI using a network pharmacology method, molecular docking, and experimental validation.

Acute kidney injury (AKI) is a severe and frequent complication of sepsis that occurs in intensive care units with inflammation and rapid decline in renal function as the main pathological features. Systemic inflammation, microvascular dysfunction, and tubule injury are the main causes of sepsis-induced AKI (SI-AKI). The high prevalence and death rate from SI-AKI is a great challenge for clinical treatment worldwide. However, in addition to hemodialysis, there is no effective drug to improve renal tissue damage and alleviate the decline in kidney function. We conducted a network pharmacological analysis of Salvia miltiorrhiza (SM), a traditional Chinese medicine, which is widely used for the treatment of kidney disease. Then, we combined molecular docking and a dynamics simulation to screen for the active monomer dehydromiltirone (DHT) that has therapeutic effects on SI-AKI and investigated its potential mechanism of action through experimental validation. The components and targets of SM were obtained by searching the database, and 32 overlapping genes were screened by intersection analysis with AKI targets. GO and KEGG data showed that the functions of a common gene were closely related to oxidative stress, mitochondrial function, and apoptosis. The molecular docking results combined with molecular dynamics simulations provide evidence for a binding model between DHT and cyclooxygenase-2 (COX2), both of which are mainly driven by van der Waals interactions and a hydrophobic effect. In vivo, we found that mice pretreated with an intraperitoneal injection of DHT (20 mg/kg/d) for 3 days ameliorated CLP surgery-induced renal function loss and renal tissue damage and inhibited inflammatory mediators IL-6, IL-1ß, TNF-α, and MCP-1 production. In vitro, the DHT pretreatment decreased LPS-induced expression of COX2, inhibited cell death and oxidative stress, alleviated mitochondrial dysfunction, and restrained apoptosis in HK-2 cells. Our research indicates that the renal preventive effect of DHT is related to maintaining mitochondrial dynamic balance, restoring mitochondrial oxidative phosphorylation, and inhibiting cell apoptosis. The findings in this study provide a theoretical basis and a novel method for the clinical therapy of SI-AKI.

Macrophage Polarization: An Important Candidate Regulator for Lung Diseases.

Macrophages are crucial components of the immune system and play a critical role in the initial defense against pathogens. They are highly heterogeneous and plastic and can be polarized into classically activated macrophages (M1) or selectively activated macrophages (M2) in response to local microenvironments. Macrophage polarization involves the regulation of multiple signaling pathways and transcription factors. Here, we focused on the origin of macrophages, the phenotype and polarization of macrophages, as well as the signaling pathways associated with macrophage polarization. We also highlighted the role of macrophage polarization in lung diseases. We intend to enhance the understanding of the functions and immunomodulatory features of macrophages. Based on our review, we believe that targeting macrophage phenotypes is a viable and promising strategy for treating lung diseases.

A DNA/DMXAA/Metal-Organic Framework Activator of Innate Immunity for Boosting Anticancer Immunity.

Immunotherapy has achieved revolutionary success in clinics, but it remains challenging for treating hepatocellular carcinoma (HCC) characterized by high vascularization. Here, it is reported that metal-organic framework-801 (MOF-801) can be employed as a stimulator of interferon genes (STING) through Toll-like receptor 4 (TLR4) not just as a drug delivery carrier. Notably, cytosine-phosphate-guanine oligodeoxynucleotides (CpG ODNs) and 5, 6-dimethylxanthenone-4-acetic acid (DMXAA) STING agonist with vascular disrupting function coordinates with MOF-801 to self-assemble into a nanoparticle (MOF-CpG-DMXAA) that effectively delivers CpG ODNs and DMXAA to cells for synergistically improving the tumor microenvironment by reprogramming tumor-associated macrophages (TAMs), promoting dendritic cells (DCs) maturation, as well as destroying tumor blood vessels. In HCC-bearing mouse models, it is demonstrated that MOF-CpG-DMXAA triggers systemic immune activation and stimulates robust tumoricidal immunity, resulting in a superior immunotherapeutic efficiency in orthotopic and recurrent HCC.

Non-crystalline light chain proximal tubulopathy associated with monoclonal gammopathy of renal significance: A case report and review of the literature.

BACKGROUND: Light chain proximal tubulopathy (LCPT) is a rare M-proteinemia-related nephropathy. Non-crystalline LCPT is even rarer. We herein report an unusual case of renal dysfunction and proteinuria due to κ-restricted and non-crystalline LCPT in a context of monoclonal gammopathy of renal significance (MGRS) without Fanconi syndrome (FS). CASE PRESENTATION: A 67-year-old man was admitted for a 2-year history of proteinuria and renal dysfunction. Fanconi syndrome (FS) was not observed. He was noted to have IgG-κ M protein, and the previous bone marrow biopsy revealed that atypical plasma cells accounted for 1.5% of the cells, which did not meet the diagnostic criteria for multiple myeloma. A renal biopsy revealed proximal tubular injury, including increased lysosomes with irregular contours and a mottled appearance without crystalline structure and the accumulation of κ light chains. He was diagnosed with non-crystalline LCPT with MGRS. Concurrently, we reviewed the non-crystalline LCPT cases previously published in the literature. Our patient finally received chemotherapy with a bortezomib and dexamethasone regimen. The patient did not seem to achieve evident nephrological and hematological remission after chemotherapy, but he was in a stable condition. CONCLUSION: Very few similar cases are reported in the literature. It is considered crucial to enhance our knowledge about these cases to establish the definition of the non-crystalline LCPT entity and allow for early diagnosis. Chemotherapy may not be necessary for all patients to maintain good renal function. Future prospective clinical research studies are necessary.

Global Epidemiology of Gaucher Disease: an Updated Systematic Review and Meta-analysis.

BACKGROUND: Gaucher disease [GD], an autosomal recessive lysosomal storage disorder, is characterized by progressive lysosomal storage of glucocerebroside in macrophages predominantly in bone, bone marrow, liver, and spleen. Meta-analysis of global GD epidemiology was not available before this study. METHODS: To provide a systematic review and meta-analysis of birth prevalence and prevalence of GD in multiple countries. MEDLINE and EMBASE databases were searched for original research articles on the epidemiology of GD from inception until July 21, 2021. Meta-analysis, adopting a random-effects logistic model, was performed to estimate the birth prevalence and prevalence of GD. RESULTS: Eighteen studies that were screened of 1874 records were included for data extraction. The studies that fulfilled the criteria for inclusion involved 15 areas/countries. The global birth prevalence of GD was 1.5 cases [95% confidence interval: 1.0 to 2.0] per 100,000 live births. The global prevalence of GD was 0.9 cases [95% confidence interval: 0.7 to 1.1] per 100,000 inhabitants. CONCLUSIONS: This is the first comprehensive systematic review that presented quantitative data of GD global epidemiology. Quantitative data on global epidemiology of GD could be the fundamental to evaluate the global efforts on building a better world for GD patients.

Preparation and Evaluation of PDMS/Carbon Soot Particles Superhydrophobic Biomimetic Composite Coating with Self-Cleaning and Durability.

In this paper, a superhydrophobic biomimetic composite coating was fabricated on brass by electrochemical etching, brushing PDMS adhesive layer, and depositing carbon soot particles. Due to the microstructure and the optimized ratio of PDMS, the contact angle of the superhydrophobic coating is up to 164° and the sliding angle is only 5°. The results of optical microscopy and morphometric laser confocal microscopy show that the prepared coating surface has a rough hierarchical structure. A high-speed digital camera recorded the droplet bouncing process on the surface of the superhydrophobic coating. The self-cleaning property of the coatings was evaluated by applying chalk dust particles as simulated solid contaminants and different kinds of liquids (including grape juice, beer, cola, and blue ink) as liquid contaminants. The coating remained superhydrophobic after physical and chemical damage tests. This work presents a strategy for fabricating superhydrophobic biomimetic composite coatings with significant self-cleaning properties, durability, and shows great potential for practical engineering applications.

Investigation of the anti-pseudorabies virus activity of interferon lambda 3 in cultured porcine kidney epithelial cells.

BACKGROUND: It has been reported Interferon-λ (IFN-λ) has stronger antiviral effect than other interferons. IFN-λ can induce antiviral interferon stimulated genes (ISGs) in epithelia to protect against virus. Pseudorabies virus (PRV) infection in pigs resulting in fatal encephalitis in newborn piglets, respiratory disorders in finishing pigs, reproductive disorders in sows and other symptoms. OBJECTIVES: Since the effect of IFN-λ on inhibiting PRV proliferation is still unknown. Inthis study, we investigate the relative contribution of porcine IFN-λ3 toward controlling the infection of PRV in vitro. Our findings may provide a new insight for the prevention and treatment of PRV. METHODS: Therefore, the antiviral assay, western blot, qRT-PCR and ELISA assay were used to investigating the contribution of IFN-λ against PRV in PK-15 cells. RESULTS: Here, we demonstrate that the replication of PRV in PK-15 cells was inhibited after pre-treatment with IFN-λ3, and such inhibition was dose dependent. Overexpression of IFN-λ3 receptor (IFNLR) also restricted virus titre in PK-15 cells. In addition, IFN-λ3 also increased the mRNA and protein expression of interferon-stimulated genes 15 (ISG15), 2'-5'-oligoadenylate synthase 1 (OAS1), IFN-inducible transmembrane 3 (IFITM3) and myxoma resistance protein 1 (Mx1) in PRV-infected PK-15 cells. Other than modulation ISGs, IFN-λ specifically activated IFN-γ mRNA expression not IFN-α or IFN-ß. CONCLUSIONS: IFN-λ3 had antiviral activity against PRV and the upregulation of ISGs and IFN-γ mRNA expression may be the mechanism of IFN-λ3's antiviral activities. Thus, IFN-λ3 has a decisive function that greatly limits PRV replication in PK-15 cells. Our study explores the antiviral activity of IFN-λ3 on PRV for the first time.

Studies on the Mechanism of the Volatile Oils from Caoguo-4 Decoction in Regulating Spleen Deficiency Diarrhea by Adjusting Intestinal Microbiota.

BACKGROUND: The Caoguo-4 decoction, a classical Mongolian medicine formula, is widely used to treat spleen deficiency diarrhea (SDD) in Mongolian for decades. Previously, the Caoguo-4 decoction volatile oil has been confirmed to be effective in ameliorating symptoms of spleen deficiency diarrhea in an animal model. However, the underlying mechanism of the Caoguo-4 decoction volatile oil is yet to be established. The aim of the current study was to investigate the antidiarrheal effects and mechanism of the Caoguo-4 decoction volatile oil. METHOD: Wistar rats were randomly divided into 5 groups of 10 animals including control, model, positive, Caoguo-4 decoction, and Caoguo-4 decoction volatile oil groups (10 rats in each group). All the rats, besides those in the control group, were induced to develop SDD by a bitter-cold purgation method with Xiaochengqi decoction. The antidiarrheal effect of Caoguo-4 decoction volatile oil was evaluated by pathological section, serum D-xylose and AMS content, plasma MTL content, and gut microbiota analysis via 16S rRNA sequencing. RESULTS: The results showed that the developed SDD rat model (model group) had decreased food intake, increased weight loss, soft stool, and bad hair color. When compared with the control group, serum was significantly reduced serum D-xylose and AML but increased MTL levels in the model group (p < 0.05). However, after treatment with either the Caoguo-4 decoction (the decoction group) or Smecta (the positive group) or volatile oil from the Caoguo-4 decoction (the volatile oil group), a significant increase in the serum D-xylose levels was observed. Additionally, AML levels significantly increased in the positive and volatile oil groups, and MTL levels significantly decreased in the decoction and volatile oil groups, when compared with the model group (p < 0.05). The pathological changes of the intestinal mucosa showed that the structure of the epithelium in the villi of the small intestine was affected, deformed, and incomplete in the model group when compared with the control group. However, either the decoction group or the volatile oil group recovered the villous morphology. The results of OTU analysis and alpha diversity analysis of intestinal bacteria showed that the intestinal microbiota of the SDD model rats showed an obvious decrease in richness and diversity of intestinal microbiota. But the intervention treatment of decoction and volatile oil could significantly recover the richness and diversity of intestinal microbiota. CONCLUSION: The intestinal microbiota destroyed in SDD modelling could be significantly improved by the Caoguo-4 decoction volatile oils, which provides reference for clinical medication.

Ultrasensitive DNA-Biomacromolecule Sensor for the Detection Application of Clinical Cancer Samples.

Diagnostic testing of biological macromolecules is of great significance for early warning of disease and cancer. Nevertheless, restricted by limited surface area and large steric hindrance, sensitive detection of macromolecules with interface-based sensing method remains challenging. Here, a "biphasic replacement" electrochemical aptamer-based (BRE-AB) sensing strategy which placed capture reaction of the biomacromolecule in a homogeneous solution phase and replaced with a small diameter of single-stranded DNA to attach to the interface is introduced. Using the BRE-AB sensor, the ultrasensitive detection of luteinizing hormone (LH) with the detection limit of 10 × 10-12 m is demonstrated. Molecular Dynamics simulations are utilized to explore the binding mechanism of aptamer and target LH. Moreover, it is confirmed that the BRE-AB sensor has excellent sensing performance in whole blood and undiluted plasma. Using the BRE-AB sensor, the LH concentrations in 40 clinical samples are successfully quantified and it is found that LH is higher expressed in breast cancer patients. Furthermore, the sensor enables simple, low-cost, and easy to regenerate and reuse, indicating potentially applicable for point-of-care biological macromolecules diagnostics.

Transdermal Administration of Volatile Oil from Citrus aurantium-Rhizoma Atractylodis Macrocephalae Alleviates Constipation in Rats by Altering Host Metabolome and Intestinal Microbiota Composition.

BACKGROUND: The Citrus aurantium- (ZhiShi, ZS-) Rhizoma Atractylodis Macrocephalae (BaiZhu, BZ) pairs are often found in herbal formulas for constipation. The volatile oils of ZS and BZ (ZBVO) have good pharmacological activity against constipation, but the mechanism for treatment of slow transit constipation (STC) remains unclear. METHOD: A rat model using diphenoxylate tablets was constructed to investigate if transdermal administration of ZBVO would mediate intestinal microorganisms and fecal metabolites and improve STC symptoms. The regulatory effects of ZBVO at 0.15, 0.30, and 0.60 mL kg-1 d-1 on STC rats were assessed by measuring fecal water content, intestinal propulsion rate, histopathology, expression of gastrointestinal hormones, brain and intestinal peptides, and inflammatory factors. The changes in intestinal flora of STC rats were analyzed by 16S rRNA gene sequencing. Moreover, the untargeted fecal metabolomics analysis was performed by ultraperformance liquid chromatography quadrupole time-of-flight mass spectrometer (UPLC-Q-TOF-MS) technology. RESULTS: The results showed that ZBVO had a modulating effect on STC by increasing the fecal water content and intestinal propulsion rate. Transdermal administration of ZBVO decreased serum levels of interleukin 6 (IL-6) and tumor necrosis factor-α (TNF-α) and increased the levels of gastrin (GAS) and substance P (SP). In addition, ZBVO increased 5-hydroxytryptamine (5-HT) levels and decreased vasoactive intestinal peptide (VIP) levels in colon and hippocampus tissues. The results of intestinal microbiota showed that ZBVO improved the diversity and abundance of intestinal microbiota and changed the community composition by decreasing Romboutsia and increasing Proteobacteria, Allobaculum, and Ruminococcaceae. And the feces metabolomics found that nicotinate and nicotinamide metabolism, purine metabolism, citrate cycle (TCA cycle), pyruvate metabolism, arachidonic acid metabolism, pyrimidine metabolism, and primary bile acid biosynthesis were modulated. CONCLUSION: These findings suggest that ZBVO can alleviate STC symptoms by promoting intestinal peristalsis, increasing fecal water content, regulating gastrointestinal hormone level, reducing the inflammatory response, and regulating brain and intestinal peptides after transdermal administration. And structural changes in the intestinal microbiota are closely related to host metabolism and intestinal microbiota destroyed in STC modeling could be significantly improved by the ZBVO, which provides a reference for the development of aromatic drug macrohealth products.

Emergence of a Salmonella enterica serovar Typhimurium ST34 isolate, CFSA629, carrying a novel mcr-1.19 variant cultured from egg in China.

OBJECTIVES: This study aimed to characterize the genomic features of a Salmonella enterica serovar Typhimurium ST34 isolate, CFSA629, which carried a novel mcr-1 variant, designated as mcr-1.19, mapped to an ESBL-encoding IncHI2 plasmid. METHODS: Antimicrobial susceptibility assays as well as WGS were carried out on isolate CFSA629. The complete closed genome was obtained and then explored to obtain genomic features. Plasmid sequence comparison was performed for pCFSA629 with similar plasmids and the mcr-1 genetic environment was analysed. RESULTS: S. Typhimurium ST34 CFSA629 expressed an MDR phenotype to six classes of compound and consisted of a single circular chromosome and one plasmid. It possessed 11 resistance genes including 2 ESBL genes that mapped to the chromosome and the plasmid; an IS26-flanked composite-like transposon was identified. A novel mcr-1 variant (mcr-1.19) was identified, which had a unique SNP (G1534A) that gave rise to a novel MCR-1 protein containing a Val512Ile amino acid substitution. Plasmid pCFSA629 possessed a conjugative plasmid transfer gene cluster as well as an antimicrobial resistance-encoding gene cluster-containing region that contained two IS26 composite-like transposonal modules, but was devoid of any plasmid-mediated quinolone resistance genes. The background of mcr-1.19 consisted of an ISApl1-mcr-1-PAP2-ter module. CONCLUSIONS: We report on an MDR S. Typhimurium ST34 CFSA629 isolate cultured from egg in China, harbouring an mcr-1.19 variant mapped to an IncHI2 plasmid. This highlights the importance of surveillance to mitigate dissemination of mcr-encoding genes among foodborne Salmonella. Improved surveillance is important for tackling the dissemination of mcr genes among foodborne Salmonella around the world.

New compounds from the stems of Fissistigma oldhamii var. longistipitatum and their cytotoxic activities.

Two new aporphine alkaloids, (R)-1,2-methylenedioxy-3,9-dimethoxy-11-hydroxy-N- carbamoyl-noraporphine (1) and 3,10,11-trimethoxy-1,2-methylenedioxy-7-oxoaporphine(2), and one new dihydrochalcone, 4',5'-dimethoxy-2'-hydroxy-3',6'-quinodihydrochalcone (3), along with five known alkaloids were isolated from the ethanol extracts of the stems of Fissistigma oldhamii var. longistipitatum. The compounds were obtained by various classical column chromatographic methods, and the structure elucidation was completed primarily on the basis of spectroscopic analysis, such as UV, NMR and HR-ESI-MS. The isolated compounds were subjected to evaluate cytotoxic activities in vitro, compound 1 had activity against HL-60 and HELA (IC50 value of 8.4 µM and 5.2 µM, respectively), compound 2 against MCF-7 (IC50 value of 3.7 µM), compound 3 against HEPG2 (IC50 value of 10.8 µM), respectively.

Clinical classification of hyperuricemia in patients with chronic kidney disease.

PURPOSE: Clinical classification of hyperuricemia (HUA) could help to guide therapy of HUA. Studies on the classification of HUA with chronic kidney disease (CKD) are rare. Therefore, we aimed to investigate the classification of HUA with CKD. METHODS: A cross-sectional study of 428 CKD patients was conducted, including 218 HUA patients. By correlation analysis, the association of 24-h urinary uric acid (24-h Uur), uric acid clearance rate (Cur), the urinary uric acid excretion per kilogram of weight per hour (Eur) and fractional excretion of uric acid (FEur) with estimated glomerular filtration rate (eGFR) was analyzed in the HUA and non-HUA groups. According to Eur combined with Cur and the 24-h Uur combined with FEur, HUA with CKD was classified into underexcretion, renal overload, combined and 'normal' types, which were also stratified by CKD stages. RESULTS: According to the Eur and Cur, in early CKD (eGFR ≥ 60 mL/min/1.73 m2), the underexcretion type accounted for 83.75%, and the renal overload type accounted for 2.5%. As the CKD stage increased, the proportion of the underexcretion type increased. According to the 24-h Uur and FEur, in early CKD, the underexcretion type accounted for 53.75%, and the renal overload type accounted for 15%. With increasing CKD stages, the proportion of the 'normal' type increased significantly. CONCLUSION: Different uses of Eur with Cur or 24-h Uur with FEur varied significantly in classifying HUA patients with CKD. Eur + Cur may be more applicable to the classification of HUA patients with CKD, and further research is needed.

AMPK alleviates high uric acid-induced Na+-K+-ATPase signaling impairment and cell injury in renal tubules.

One of the mechanisms in hyperuricemia (HUA)-induced renal tubular injury is the impairment of Na+-K+-ATPase (NKA) signaling, which further triggers inflammation, autophagy, and mitochondrial dysfunction and leads to cell injury. Here, we used RNA sequencing to screen the most likely regulators of NKA signaling and found that the liver kinase B1(LKB1)/adenosine monophosphate (AMP)-activated protein kinase (AMPK)/ mammalian target of rapamycin (mTOR) pathway was the most abundantly enriched pathway in HUA. AMPK is a key regulator of cell energy metabolism; hence, we examined the effect of AMPK on HUA-induced dysregulation of NKA signaling and cell injury. We first detected AMPK activation in high uric acid (UA)-stimulated proximal tubular epithelial cells (PTECs). We further found that sustained treatment with the AMPK activator 5-aminoimidazole-4-carboxamide 1-ß-d-ribofuranoside (AICAR), but not the AMPK inhibitor Compound C, significantly alleviated UA-induced reductions in NKA activity and NKA α1 subunit expression on the cell membrane by reducing NKA degradation in lysosomes; sustained AICAR treatment also significantly alleviated activation of the NKA downstream molecules Src and interleukin-1ß (IL-1ß) in PTECs. AICAR further alleviated high UA-induced apoptosis, autophagy, and mitochondrial dysfunction. Although AMPK activation by metformin did not reduce serum UA levels in hyperuricemic rats, it significantly alleviated HUA-induced renal tubular injury and NKA signaling impairment in vivo with effects similar to those of febuxostat. Our study suggests that AMPK activation may temporarily compensate for HUA-induced renal injury. Sustained AMPK activation could reduce lysosomal NKA degradation and maintain NKA function, thus alleviating NKA downstream inflammation and protecting tubular cells from high UA-induced renal tubular injury.

Adeno-associated Virus 9-mediated Small RNA Interference of TLR4 Alleviates Myocardial Ischemia and Reperfusion Injury by Inhibition of the NF-κB and MAPK Signaling Pathways in Rats.

BACKGROUND: Despite intensive investigation, effective therapeutic procedures for myocardial I/R injury are still in demand. OBJECTIVE: To explore the effect of adeno-associated virus 9 (AAV9)-mediated small interfering RNA targeting TLR4 in the treatment of myocardial ischemia and reperfusion (I/R) injury and its influence on the NF-κB and MAPK signaling pathways. METHODS: Rats were divided into 3 groups, namely, the sham, AAV9-siRNA control, and AAV9-TLR4 siRNA groups. siRNA solution or normal saline was injected through the tail vain. The rat myocardial I/R injury model was then established. HE staining and TUNEL staining were applied to compare the pathological changes in cardiomyocytes in the three groups. Immunohistochemical staining and western blotting were utilized to detect TLR4 expression under siRNA interference. Serum inflammatory factor (IL-1ß, TNF-α) expression was determined by an ELISA commercial kit. Key proteins in the MAPK (p38, JNK 1/2) and NF-κB (p65) signaling pathways were determined to identify the TLR4 siRNA functional mechanism. RESULTS: Fluorescence microscopic images of the myocardium indicated that AAV9- mediated siRNA was efficiently transfected into the myocardium, and the infarcted size after I/R injury was decreased by AAV9-TLR4 siRNA when compared with negative control rats (P<0.05). TLR4 protein expression was significantly decreased by siRNA interference (P<0.001). Apoptosis-related factor BCL-2 expression was increased in the TLR4 gene silencing group, whereas Bax expression was decreased. The Bax/BCL-2 ratio was also decreased, demonstrating a protective effect for cardiomyocytes. Inflammatory factors were lower in the TLR4 gene silencing group than in the siRNA control group (P<0.001). The MAPK and NF-κB signaling pathways were activated in myocardial I/R injury; however, the primary proteins in these two signaling pathways were downregulated upon interference of TLR4 siRNA, with significant differences (P<0.05). CONCLUSION: AAV9-TLR4 siRNA has a positive effect on myocardial I/R injury by inhibiting the MAPK and NF-κB signaling pathways and can be used as a potential therapeutic method for myocardial I/R injury.