Systemic and Local Administration of a Dual-siRNA Complex Efficiently Inhibits Tumor Growth and Bone Invasion in Oral Squamous Cell Carcinoma.

Oral squamous cell carcinoma (OSCC) accounts for nearly 90% of oral and oropharyngeal cancer cases and is characterized by high mortality and poor prognosis. RNA-based gene therapies have been developed as an emerging option for cancer treatment, but it has not been widely explored in OSCC. In this work, we developed an efficient siRNA cationic micelle DOTAP-mPEG-PCL (DMP) by self-assembling the cationic lipid DOTAP and monomethoxy poly(ethylene glycol)-poly(ε-caprolactone) (mPEG-PCL) polymer. We tested the characteristics and transformation efficiency of this micelle and combined DMP with siRNA targeting STAT3 and TGF-ß to evaluate the antitumor effect and bone invasion interfering in vitro and in vivo. The average size of the DMP was 28.27 ± 1.62 nm with an average zeta potential of 54.60 ± 0.29 mV. The DMP/siRNA complex showed high delivery efficiency, with rates of 97.47 ± 0.42% for HSC-3. In vitro, the DMP/siSTAT3 complex exhibited an obvious cell growth inhibition effect detected by MTT assay (an average cell viability of 25.1%) and clonogenic assay (an average inhibition rate of 51.9%). Besides, the supernatant from HSC-3 transfected by DMP/siTGF-ß complexes was found to interfere with osteoclast differentiation in vitro. Irrespective of local or systemic administration, DMP/siSTAT3+siTGF-ß showed antitumor effects and bone invasion inhibition in the OSCC mice mandibular invasion model according to tumor volume assays and Micro-CT scanning. The complex constructed by DMP cationic micelles and siSTAT3+siTGF-ß represents a potential RNA-based gene therapy delivery system for OSCC.

Clinical activity and safety of sintilimab, bevacizumab, and TMZ in patients with recurrent glioblastoma.

PURPOSE: There are limited and no standard therapies for recurrent glioblastoma. We herein report the antitumour activity and safety of sintilimab, bevacizumab and temozolomide (TMZ) in recurrent glioblastoma. METHODS: We retrospectively analysed eight patients with recurrent glioblastoma treated with sintilimab (200 mg) every three weeks + bevacizumab (10 mg/kg) every three weeks + TMZ (200 mg/m²orally) (5 days orally every 28 days for a total of four weeks). The primary objective was investigator-assessed median progression-free survival(mPFS). Secondary objectives were to assess the 6-month PFS, objective response rate (ORR) and duration of response (DOR) accroding to RANO criteria. RESULTS: The mPFS time for 8 patients was 3.340 months (95% CI: 2.217-4.463), The longest PFS was close to 9 months. Five patients were assessed to have achieved partial response (PR), with an overall remission rate of 62.5%, Four patients experienced a change in tumour volume at the best response time of greater than 60% shrinkage from baseline, and one patient remained progression free upon review, with a DOR of more than 6.57 months. The 6-month PFS was 25% (95% CI: 5.0-55.0%). Three patients had a treatment-related adverse events, though no grade 4 or 5 adverse events occurred. CONCLUSION: In this small retrospective study, the combination regimen of sintilimab, bevacizumab and TMZ showed promising antitumour activity in treatment of recurrent glioblastoma, with a good objective remission rate.

Injectable and Photocurable Gene Scaffold Facilitates Efficient Repair of Spinal Cord Injury.

RNA interference-based gene therapy has led to a strategy for spinal cord injury (SCI) therapy. However, there have been high requirements regarding the optimal gene delivery vector for siRNA-based SCI gene therapy. Here, we developed an injectable and photocurable lipid nanoparticle GelMA (PLNG) hydrogel scaffold for controlled dual siRNA delivery at the SCI wound site. The prepared PLNG scaffold could efficiently protect and retain the bioactivity of the siRNA nanocomplex. It facilitated sustainable siRNA release along with degradation in 7 days. After loading dual siRNA targeting phosphatase and tensin homologue (PTEN) and macrophage migration inhibitory factor (MIF) simultaneously, the locally administered siRNAs/PLNG scaffold efficiently improved the Basso mouse scale (BMS) score and recovered ankle joint movement and plantar stepping after treatment with only three doses. We further proved that the siRNAs/PLNG scaffold successfully regulated the activities of neurons, microglia, and macrophages, thus promoting neuron axon regeneration and remyelination. The protein array results suggested that the siRNAs/PLNG scaffold could increase the expression of growth factors and decrease the expression of inflammatory factors to regulate neuroinflammation in SCI and create a neural repair environment. Our results suggested that the PLNG scaffold siRNA delivery system is a potential candidate for siRNA-based SCI therapy.

Establishment of an efficient callus transient transformation system for Vitis vinifera cv. 'Chardonnay'.

Grape (Vitis L.), a highly valued fruit crop, poses significant challenges in genetic transformation and functional characterization of genes. Therefore, there is an urgent need for the development of a rapid and effective method for grape transformation and gene function identification. Here, we introduce a streamlined Agrobacterium-mediated transient transformation system for grape calli. Optimal conditions were established with a leaf-derived callus induction medium; chiefly B5 medium supplemented with 0.05 mg/L NAA, 0.5 mg/L 2,4-D, and 2.0 mg/L KT; and a callus proliferation medium (B5 medium supplemented with 0.5 mg/L NAA and 2.0 mg/L 6-BA), respectively. Notably, GUS enzyme activity peaked (352.96 ± 33.95 mol 4-MU/mg/min) by sonication with Agrobacterium tumefaciens EHA105 and 100 µM AS for 4 min, followed by vacuum infection for 5 min, and co-culture at 25 °C in the dark for 1 day using callus as explants at an optical density (OD600) of 0.8. VaCIPK18 gene was transiently transformed into calli, and transcripts of the gene (endogenous and exogenous) were detected at higher levels than in non-transformed calli (endogenous). Moreover, after 10 days of treatment at 4 °C or -4 °C, the callus net weight of transformed callus was significantly higher than that of the untransformed callus, indicating that the VaCIPK18-overexpressing grape callus could improve cold tolerance. Overall, we establish a simple but effective transient transformation approach for grape callus, which could serve as a useful tool for the rapid assessment of gene function in this important crop.

Tumor Cell Lysate-Based Multifunctional Nanoparticles Facilitate Enhanced mRNA Delivery and Immune Stimulation for Melanoma Gene Therapy.

Messenger ribonucleic acid (mRNA)-based gene therapy has great potential for cancer gene therapy. However, the effectiveness of mRNA in cancer therapy needs to be further improved, and the delivery efficiency and instability of mRNA limit the application of mRNA-based products. Both the delivery efficiency can be elevated by cell-penetrating peptide modification, and the immune response can be enhanced by tumor cell lysate stimulation, representing an advantageous strategy to expand the effectiveness of mRNA gene therapy. Therefore, it is vital to exploit a vector that can deliver high-efficiency mRNA with codelivery of tumor cell lysate to induce specific immune responses. We previously reported that DMP cationic nanoparticles, formed by the self-assembly of DOTAP and mPEG-PCL, can deliver different types of nucleic acids. DMP has been successfully applied in gene therapy research for various tumor types. Here, we encapsulated tumor cell lysates with DMP nanoparticles and then modified them with a fused cell-penetrating peptide (TAT-iRGD) to form an MLSV system. The MLSV system was loaded with encoded Bim mRNA, forming the MLSV/Bim complex. The average size of the synthesized MLSV was 191.4 nm, with a potential of 47.8 mV. The MLSV/mRNA complex promotes mRNA absorption through caveolin-mediated endocytosis, with a transfection rate of up to 68.6% in B16 cells. The MLSV system could also induce the maturation and activation of dendritic cells, obviously promoting the expression of CD80, CD86, and MHC-II both in vitro and in vivo. By loading the encoding Bim mRNA, the MLSV/Bim complex can inhibit cell proliferation and tumor growth, with inhibition rates of up to 87.3% in vitro. Similarly, the MLSV/Bim complex can inhibit tumor growth in vivo, with inhibition rates of up to 78.7% in the B16 subcutaneous tumor model and 63.3% in the B16 pulmonary metastatic tumor model. Our results suggest that the MLSV system is an advanced candidate for mRNA-based immunogene therapy.

Exogenous Melatonin Promotes Cold Tolerance in Grape Seedlings: Physiological, Transcriptomic, and Functional Evidence.

Melatonin (MEL) is an antioxidant molecule that enhances plant tolerance to environmental stress. However, the mechanisms by which MEL regulates cold signaling pathways in grapes under cold stress remain elusive. Here, we investigated the physiological and transcriptomic changes in grape seedlings treated with exogenous MEL to determine their protective role under cold stress. Results showed that 150 µM MEL effectively attenuated cold-induced cell damage by reducing reactive oxygen species (ROS) and preserving the chloroplast structure and function. MEL also inhibited tannin degradation, which contributed to its protective effect. Exogenous MEL promoted the synthesis of endogenous MEL, abscisic acid, auxin, and cytokinin while inhibiting gibberellin. Transcriptomic profiling revealed 776 differentially expressed transcripts in MEL-treated samples compared to controls. Functional analysis of a candidate hub gene, VvHSFA6b, showed that its overexpression in grape calli enhances cold tolerance by activating jasmonic acid synthesis pathway genes, promoting JA accumulation, and inhibiting JAZ-repressed transcription factors.

Targeted siRNA Delivery by Bioinspired Cancer Cell Membrane-Coated Nanoparticles with Enhanced Anti-Cancer Immunity.

Introduction: Cell-membrane nanocarriers are usually constructed by modifying the nanoparticle surface with cell membrane extracts, which has a direct benefit in endowing targeting capacity to nanocarriers based on their original cell types. However, delivering nucleic acid cargos by cell membrane-based nanoparticles is difficult owing to the strong negative charge of the cell membrane fraction. In this study, we developed a cancer cell membrane-based drug delivery system, the cMDS, for efficient siRNA delivery. Meanwhile, the cancer-specific immune response stimulated by the gene vector itself could offer synergistic anti-cancer ability. Methods: The cMDS was prepared by ultrasound, and its transfection efficiency and anti-cancer ability were examined using cultures of CT26 cells. MTT and red blood cell hemolysis tests were performed to assess the safety of cMDS, while its targeted gene delivery and strong immune stimulation were investigated in a subcutaneous tumor model. Moreover, the detailed anti-cancer immune stimulation mechanisms of cMDS are uncovered by protein chip analysis. Results: The cMDS was spherical core-shell structure. It showed high transfection efficiency and anti-cancer ability in vitro. In animal experiments, intravenously administered cMDS/siStat3 complex efficiently suppress the growth of colon cancer. Moreover, the result of protein chip analysis suggested that cMDS affect the migration and chemotaxis of immune cells. Conclusion: The cMDS shows obvious tumor tissue-specific accumulation properties and strong immune stimulation ability. It is an advanced targeted gene delivery system with potent immunotherapeutic properties.

Defects Detection Method Based on Programmable Spoof Surface Plasmon Polaritons in Non-Metallic Composites.

Microwave nondestructive testing (NDT) offers promising application prospects due to its advantages of non-contact inspection in detecting defects in non-metallic composites. However, the detection sensitivity of this technology is generally affected by the lift-off effect. To reduce this effect and highly concentrate electromagnetic fields on defects, a defect detection method using scanning instead of moving sensors in the microwave frequency range was proposed. Additionally, a novel sensor based on the programmable spoof surface plasmon polaritons (SSPPs) was designed for non-destructive detection in non-metallic composites. The unit structure of the sensor was made up of a metallic strip and a split ring resonator (SRR). A varactor diode was loaded between the inner and outer rings of the SRR, and by changing the capacitance of this diode using electronic scanning, the field concentration phenomenon of the SSPPs sensor can be moved along a specific direction for defect detection. By using this proposed method and sensor, the location of a defect can be analyzed without moving the sensor. The experimental results demonstrated that the proposed method and designed SSPPs sensor can be effectively applied in detecting defects in non-metallic materials.

Non-mechanical self-alignment system for free-space optical communications based on a cascaded liquid crystal optical antenna.

Free-space optical (FSO) communication has attracted extensive attention in recent years. To maintain a reliable FSO link, two main issues need to be addressed: beam drift and vibration. In this paper, we demonstrate a non-mechanical self-alignment system based on a cascaded liquid crystal optical antenna, in which a frequency decoupled hybrid integration Kalman filter (FDHI-KF) method is proposed to achieve predictive beam drift tracking and vibration mitigation. By leveraging the integrated control on our lab-made liquid crystal phase modulation devices, and implementing the adaptive algorithm on a heterogeneous field programmable gate array (FPGA), this system is capable of realizing precise self-alignment without any moving parts. Experiments are conducted to verify its performance in practical applications. We envision it to set a benchmark for future liquid crystal non-mechanical beam-steering systems in FSO communications.

Small molecules in the treatment of COVID-19.

The outbreak of COVID-19 has become a global crisis, and brought severe disruptions to societies and economies. Until now, effective therapeutics against COVID-19 are in high demand. Along with our improved understanding of the structure, function, and pathogenic process of SARS-CoV-2, many small molecules with potential anti-COVID-19 effects have been developed. So far, several antiviral strategies were explored. Besides directly inhibition of viral proteins such as RdRp and Mpro, interference of host enzymes including ACE2 and proteases, and blocking relevant immunoregulatory pathways represented by JAK/STAT, BTK, NF-κB, and NLRP3 pathways, are regarded feasible in drug development. The development of small molecules to treat COVID-19 has been achieved by several strategies, including computer-aided lead compound design and screening, natural product discovery, drug repurposing, and combination therapy. Several small molecules representative by remdesivir and paxlovid have been proved or authorized emergency use in many countries. And many candidates have entered clinical-trial stage. Nevertheless, due to the epidemiological features and variability issues of SARS-CoV-2, it is necessary to continue exploring novel strategies against COVID-19. This review discusses the current findings in the development of small molecules for COVID-19 treatment. Moreover, their detailed mechanism of action, chemical structures, and preclinical and clinical efficacies are discussed.

Local and Systemic Delivery of the BimS Gene Nano-Complex for Efficient Oral Squamous Cell Carcinoma Therapy.

Purpose: Oral squamous cell carcinoma (OSCC) is the most common type of oral cancer, with more than 300,000 new cases annually. Despite advances in existing treatments, including surgery, radiation, chemotherapy, and immunotherapy, the overall survival and prognosis have remained poor. However, gene therapy based on non-viral vectors provides new ideas for the treatment of OSCC. Here, we aimed to prepare and describe the synthesis, biosafety, and preclinical efficacy of DOTAP-mPEG-PCL (DMP) in OSCC gene therapy. Methods: We prepared a nano-sized hybrid cationic micelle DMP. DMP micelles were prepared by self-assembling cationic lipid DOTAP and mPEG-PCL polymer. We evaluated the characteristics of this cationic micelle in vitro. Combined with encoding the apoptosis-inducing BimS gene, we established the DMP/phBimS complex and evaluated its anti-tumor effect in vitro. We also established a mouse tongue xenograft model to evaluate the antitumor effect of the DMP/phBimS complex in vivo through local and systemic administration prospectively. Results: The DMP cationic micelle is spherical in shape, with an average diameter of 28.32 ± 3.56 nm and an average zeta potential of 43.43 ± 0.82 mV. By activation of lipid raft-mediated endocytosis caveolin-mediated endocytosis, DMP could efficiently deliver plasmid into SCC15 cells (efficiency: 52.07% ± 1.63%), with an ideal biosecurity. When loaded by plasmid encoding the apoptosis-inducing BimS gene, the DMP/phBimS complex exhibited an obvious anti-proliferation effect of SCC15 in vitro through the apoptosis pathway (33.9% ± 2.62% apoptosis rate). By local administration, the DMP/phBimS complex showed ideal anti-tumor properties in the nude mouse tongue xenograft model, with an average tumor inhibition rate of 65.66%. Furthermore, through systematic administration, the DMP/phBimS complex obviously inhibited OSCC growth, with an average inhibition rate of 45.63% (DMP/phBimS) and an appropriate biocompatibility. Conclusion: The DMP/phBimS complex is an optional effective option for suicide gene therapy for OSCC.

Dual-RNA controlled delivery system inhibited tumor growth by apoptosis induction and TME activation.

Drug-controlled release is recognized as effective for improving compliance with treatment and obtaining better therapeutic efficacy with less toxicity in cancer treatment. However, few reports in this area are involved in nucleic acids delivery, especially in RNA therapeutics delivery. In this study, an injectable hydrogel Methacrylated gelatin (GM) scaffold was introduced into a dual-RNA hybrid delivery complex hybrid lipid particle (HLP) to form a G-HLP/RNAs system. This system can control the release of both siRNA and mRNA and was found to be efficient for protecting these RNAs from biodegradation and retaining their therapeutic effect over 7 days. Further, a tumor environment (TME)-activation function after peritumoral injection of mocked GM scaffold was observed. Then, matured DC cells and activated T-cells were detected by the addition of HLP/RNAs complex, thus verifying the immunoactivation function of GM scaffold and its ability to reserve immune cells and antigens. Finally, two doses of G-HLP/RNAs treatment efficiently suppressed C26 tumor growth in mice with a tumor weight inhibition rate of 71.9%. Owing to its ability to achieve RNA drug-controlled release, alter TME, and induce tumor apoptosis, the G-HLP/RNAs system may become a valuable tool for cancer gene therapy.

Efficient Treatment of Rheumatoid Arthritis by Degradable LPCE Nano-Conjugate-Delivered p65 siRNA.

Rheumatoid arthritis (RA) is one of the most common autoimmune diseases worldwide, causing severe cartilage damage and disability. Despite the recent progress made in RA treatment, limitations remain in achieving early and efficient therapeutic intervention. Advanced therapeutic strategies are in high demand, and siRNA-based therapeutic technology with a gene-silencing ability represents a new approach for RA treatment. In this study, we created a cationic delivery micelle consisting of low-molecular-weight (LMW) polyethylenimine (PEI)-cholesterol-polyethylene glycol (PEG) (LPCE) for small interfering RNA (siRNA)-based RA gene therapy. The carrier is based on LMW PEI and modified with cholesterol and PEG. With these two modifications, the LPCE micelle becomes multifunctional, and it efficiently delivered siRNA to macrophages with a high efficiency greater than 70%. The synthesized LPCE exhibits strong siRNA protection ability and high safety. By delivering nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) p65 siRNA, the p65 siRNA/LPCE complex efficiently inhibited macrophage-based cytokine release in vitro. Local administration of the p65 siRNA/LPCE complex exhibited a fast and potent anti-inflammatory effect against RA in a mouse model. According to the results of this study, the functionalized LPCE micelle that we prepared has potential gene therapeutic implications for RA.

Single Micelle Vectors based on Lipid/Block Copolymer Compositions as mRNA Formulations for Efficient Cancer Immunogene Therapy.

Immunogene therapy provides a new strategy for the treatment of colorectal cancer. Compared to plasmid DNA, mRNA possesses several advantages as a therapeutic nucleic acid material and shows high potential in cancer therapy. Although efforts have been made to conquer the limited efficiency of mRNA delivery, most of the current mRNA vectors possess complex structures or compositions, which introduces additional toxicity and hinders their further clinical application. Hence, it is highly necessary to develop potent mRNA delivery systems with simple structures. Here, we report efficient mRNA delivery using the biodegradable micelle delivery system of DMP (DOTAP-mPEG-PCL). Biodegradable DMP micelles were simply prepared by the self-assembly of cationic lipid DOTAP and the diblock polymer monomethoxy poly(ethylene glycol)-poly(ε-caprolactone). With an average size of only 30 nm, we proved that these single-structured cationic micelles are highly potent in condensing and protecting mRNA molecules, with a delivery efficiency of 60.59% on C26 mouse colon cancer cells. The micelles triggered specific internalization pathways and were fully degraded in vivo. After binding with IL-22BP (interleukin-22 binding protein)-encoding mRNA, a strongly elevated IL-22BP mRNA level was detected in C26 cells. After intraperitoneal and intratumoral injection of the DMP/mIL-22BP complex, strong inhibition effects on C26 colon cancer models were observed, with high therapeutic efficiency and safety when systemically administrated. These data suggest that the DMP micelle is an advanced single-structured mRNA delivery system with high safety.

Functionalized DMP-039 Hybrid Nanoparticle as a Novel mRNA Vector for Efficient Cancer Suicide Gene Therapy.

BACKGROUND: Gene therapy has emerged as a new strategy for cancer therapy. As an alternative nucleic acid material, messenger ribonucleic acid (mRNA) is being increasingly utilized in cancer gene therapy. However, unfulfilled requirements and a lack of ideal mRNA delivery vectors persist. METHODS: We developed an advanced mRNA delivery system, DMP-039, by fusing a cell-penetrating peptide, cRGD-R9, and a cationic nano-sized DMP backbone together. The DMP gene vector backbone was synthesized by the self-assembly of DOTAP lipid and mPEG-PCL polymer. Introduction of the cRGD-R9 peptide onto the DMP backbone was performed to elevate the mRNA delivery capacity, which resulted in a peptide-functionalized hybrid delivery system. RESULTS: The average size of the synthesized DMP-039 was 268.9 ± 12.4 nm (PDI = 0.382), with a potential of 17.4 ± 0.5 mV. The synthesized DMP-039 hybrid nanoparticles exhibited high mRNA delivery efficiency through multiple mechanisms during transmembrane transportation. By loading the encoding mRNA from the suicide gene Bim, a locally administered mBim/DMP-039 complex strongly inhibited growth in two colon cancer models. Moreover, intravenous administration of the mBim/DMP-039 complex efficiently suppressed C26 pulmonary metastatic tumor progression with high safety. The in vivo distribution, degradation, and excretion were also investigated in detail. CONCLUSION: Our results suggest that the DMP-039 peptide-functionalized hybrid nanoparticle is an advanced candidate for mRNA-based suicide gene therapy.

Current Progress in Messenger RNA-Based Gene Therapy.

Based on its rapid expression, simple sequence composition, low immunogenicity, and flexible modification possibilities, in vitro synthesized mRNA has demonstrated strong potential as a candidate for gene therapy. Many efforts have been made to enhance its therapeutic efficacy and safety. Profiting from the development in pathogenesis and materials science, much progress has been achieved in mRNA-based therapy studies. Many mRNA-derived therapeutics including vaccines, antibodies, cytokines, and growth factors have emerged for the treatment of diverse diseases that have multiple modes of action. Novel delivery vectors with enhanced capacity, safety, and properties have been developed to meet the demands of mRNA delivery. Advanced strategies like library screening, environment interaction, and bio-inspiration materials have been used in the investigation process and produced valuable results. In this review, we summarize and discuss recent advances in mRNA-based gene therapy studies.

Distribution law of dust concentration by image transmission in a cement workshop.

Research on the distribution law of dust concentration is conducive to controlling dust pollution in industry workshops. The traditional experimental method has a common disadvantage in that it only can measure dust concentration of a few sampling points, resulting in limitation of further research about the distribution law of dust concentration in cement workshops. Aimed at revealing more visually and accurately the distribution law of dust concentration, a vision-based measurement (VBM) system for dust concentration based on the image transmission is constructed by utilizing an established dust simulation device in cement workshops. The measurement system measured the dust concentration in the X and Z directions with a line-scanning measuring mode and that of end face XOZ in a face-scanning measuring mode. Experimental results show that the distribution law of dust concentration can be regarded as a first exponential function and then oscillating function relationship between the dust concentration and the distance from the dust source in the X direction, a parabolic function between the dust concentration and the distance from the dust source in the Z direction, and a type of laminar or turbulence distribution in the end face XOZ. The proposed VBM and the revealed law can provide a reference for the study of dust concentration distribution and dust control.

[Effects of telmisartan on the expression of adiponectin and its receptors in testes of type 1 diabetic rats].

OBJECTIVE: To explore the effects of telmisartan on the expressions of adiponectin and adiponectin receptor and its signal transduction pathway AMP-activated protein kinase (AMPK), Akt/endothelial nitric oxide synthase (Akt/e-NOS/NO) and examine the possible protective mechanisms of telmisartan in testis of type 1 diabetic rats. METHODS: A total of 27 male Sprague-Dawley rats were randomly divided into normal control (NC, n=8) group and diabetic model (n=19) group. Diabetes was induced by an intraperitoneal injection of streptozotocin (STZ). And 16 established diabetic rats were randomly divided into diabetic control (DM, n=8) and diabetic treated with telmisartan (DT, n=8) groups. Group DT received a lavage of telmisartan while groups NC and DM had an equal volume of normal saline by gavage. At the end of 8-week of telmisartan treatment, the animals were sacrificed after harvesting of blood samples. Bilateral testes were extracted and epididymis was minced for preparing sperm suspension. Blood samples were used to detect the related parameters. Some tresticular tissues were fixed in neutral 40% formaldehyde solution and processed for histological analysis. And the remaining testicular tissues were immediately placed into liquid nitrogen and then stored in a -70 °C refrigerator until analyses. The levels of insulin and sex hormone were detected by radioimmunoassay. The levels of adiponectin, interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) were assayed by enzyme-linked immunosorbent assay (ELISA). The mRNA expression of testicular adiponectin and its receptor was assessed by real-time fluorescent quantitative polymerase chain reaction (PCR). The protein expressions of testicular adiponectin, adiponectin receptor, AMPK, P-AMPK, AKT, P-AKT and e-NOS were analyzed by Western blot. RESULTS: There was significant pathological changes in testes in group DM than that in group NC. The levels of P-AKT/AKT, e-NOS and NO were significantly increased in group DM than those in group NC [(1.54 ± 0.27) vs (1.00 ± 0.00), (1.56 ± 0.26) vs (1.00 ± 0.00), (1.75 ± 0.28) vs (1.08 ± 0.02) µmol/g, all P<0.05]. The levels of serum adiponectin,testicular adiponectin and its receptor 1, and the ratio of P-AMPK to AMPK significantly decreased in group DM than that in group NC [(622.46 ± 95.86) vs (2 022.07 ± 51.13)ng/ml, (0.66 ± 0.09) vs (1.00 ± 0.00), (0.68 ± 0.05) vs (1.00 ± 0.00), (0.34 ± 0.11) vs (1.00 ± 0.00), all P<0.05]. There was no significant difference in the expression of adiponectin receptor 2 between group DM and NC [(1.02 ± 0.13) vs (1.00 ± 0.00), P>0.05]. After 8-week telmisartan treatment, the pathological changes of testes became alleviated in diabetic rats. The levels of P-AKT/AKT, e-NOS and NO significantly decreased in group DT than those in group DM [(1.24 ± 0.39) vs (1.54 ± 0.27), (1.16 ± 0.47) vs (1.56 ± 0.26), (1.35 ± 0.30) vs (1.75 ± 0.28) µmol/g, all P<0.05]. The serum and testicular levels of adiponectin and testicular adiponectin receptor 1 significantly increased in group DT than those in group DM [(1 051.55 ± 102.55) vs (622.46 ± 95.86), (0.84 ± 0.09) vs (0.66 ± 0.09), (0.80 ± 0.07) vs (0.68 ± 0.05), all P<0.05]. No significant difference existed in the ratio of P-AMPK to AMPK in testes or the expression of adiponectin receptor 2 between group DM and NC [(0.65 ± 0.52) vs (0.34 ± 0.11), (1.02 ± 0.15) vs (1.02 ± 0.13), all P>0.05]. CONCLUSION: Telmisartan may reduce the injury degree of testes and play protective roles in testicular tissues in diabetic rats by regulating the expressions of adiponectin, adiponectin receptor and the signal pathways mediated by adiponectin receptor.

Effect of exenatide on the cardiac expression of adiponectin receptor 1 and NADPH oxidase subunits and heart function in streptozotocin-induced diabetic rats.

BACKGROUND: This study investigated the effect of exenatide on the cardiac expression of adiponectin receptor 1 and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits and heart function in streptozotocin-induced diabetic rats. METHODS: Male Sprague-Dawley rats were randomly divided into four groups, i.e. control group, diabetic group, diabetic treated with low doses of exenatide (2 µg · kg-1.d-1) and diabetic treated with high doses of exenatide (10 µg · kg-1.d-1). Diabetes was induced by intraperitoneal injection of streptozotocin (65 mg/kg body weight). At the termination after exenatide treatment for eight weeks, following anesthesia of the rats, a catheter was inserted into the left ventricle through the right common carotid artery for measurement of left ventricular pressure, which included left ventricular systolic pressure (LVSP), left ventricular end-diastolic pressure (LVEDP) and the maximal rate of rise and decline of ventricular pressure (±dp/dt[max]). Plasma and myocardial adiponectin levels, and the expressions of myocardial adiponectin receptor 1, p22phox, NADPH oxidase 4 (NOX4), glucose transporter type 4 (Glut4), AMPK-α, phosphorylated-AMPK-α, connective tissue growth factor (CTGF) and copper zinc superoxide dismutase (Cu-Zn-SOD) were assayed. RESULTS: Heart function, plasma adiponectin levels, the protein expression of myocardial phosphorylated-AMPK-α, the mRNA expression of myocardial Glut4, and the positive expression of myocardial Cu-Zn-SOD were significantly decreased in diabetic. The protein expression of myocardial adiponectin receptor 1, the mRNA expression of myocardial p22phox and NOX4, and the positive expression of myocardial CTGF were significantly increased in diabetic. Low and high doses of exenatide treatment significantly attenuated these changes in diabetic rats. CONCLUSIONS: These results suggest that exenatide may contribute to the improvement of the heart function in diabetic rats by down-regulating the expression of myocardial adiponectin receptor 1, p22phox and NOX4, and up-regulating plasma adiponectin level and the expression of myocardial AMPK-α, Glut4 and Cu-Zn-SOD.

Peroxisome proliferator-activated receptor gamma ligands induce cell cycle arrest and apoptosis in human renal carcinoma cell lines.

AIM: To study the effect of peroxisome proliferator-activated receptor gamma (PPARgamma) ligands on cell proliferation and apoptosis in human renal carcinoma cell lines. METHODS: The expression of PPARgamma was investigated by reverse transcriptase polymerase chain reaction (RT-PCR), Western blot and immunohistochemistry. The effect of thiazolidinedione (TZD) PPARgamma ligands on growth of renal cell carcinoma (RCC) cells was measured by MTT assay and flow cytometric analysis. Cell death ELISA, Hoechst 33342 fluorescent staining and DNA ladder assay were used to observe the effects of PPAR gamma ligands on apoptosis. Regulatory proteins of cell cycle and apoptosis were detected by Western blot analysis. RESULTS: PPARgamma was expressed at much higher levels in renal tumors than in the normal kidney (2.16+/-0.85 vs 0.90+/-0.73; P<0.01). TZD PPARgamma ligands inhibited RCC cell growth in a dose-dependent manner with IC50 values of 7.08 micromol/L and 11.32 micromol/L for pioglitazone, and 5.71 micromol/L and 8.38 micromol/L for troglitazone in 786-O and A498 cells, respectively. Cell cycle analysis showed a G0/G1 arrest in human RCC cells following 24-h exposure to TZD. Analysis of cell cycle regulatory proteins revealed that TZD decreased the protein levels of proliferating cell nuclear antigen, pRb, cyclin D1, and Cdk4 but increased the levels of p21 and p27 in a time-dependent manner. Furthermore, high doses of TZD induced massive apoptosis in renal cancer cells, with increased Bax expression and decreased Bcl-2 expression. CONCLUSION: TZD PPAR gamma ligands showed potent inhibitory effect on proliferation, and could induce apoptosis in RCC cells. These results suggest that ligands for PPAR gamma have potential antitumor effects on renal carcinoma cells.