Neuropathic pain development following nerve injury is mediated by SOX11-ARID1A-SOCS3 transcriptional regulation in the spinal cord.

BACKGROUND: Neuropathic pain, a complex condition originating from nervous system damage, remains a significant clinical challenge due to limited understanding of its underlying mechanisms. Recent research highlights the SOX11 transcription factor, known for its role in nervous system development, as a crucial player in neuropathic pain development and maintenance. This study investigates the role of the SOX11-ARID1A-SOCS3 pathway in neuropathic pain modulation within the spinal cord. METHODS AND RESULTS: Using a spinal nerve ligation (SNL) model in mice, we observed a significant upregulation of Sox11 in the spinal cord dorsal horn post-injury. Intrathecal administration of Sox11 shRNA mitigated SNL-induced neuropathic pain behaviors, including mechanical allodynia and heat hyperalgesia. Further, we demonstrated that Sox11 regulates neuropathic pain via transcriptional control of ARID1A, with subsequent modulation of SOCS3 expression. Knockdown of ARID1A and SOCS3 via shRNA resulted in alleviation of Sox11-induced pain sensitization. Additionally, Sox11 overexpression led to an increase in ARID1A binding to the SOCS3 promoter, enhancing chromatin accessibility and indicating a direct regulatory relationship. These findings were further supported by in vitro luciferase reporter assays and chromatin accessibility analysis. CONCLUSIONS: The SOX11-ARID1A-SOCS3 pathway plays a pivotal role in the development and maintenance of neuropathic pain. Sox11 acts as a master regulator, modulating ARID1A, which in turn influences SOCS3 expression, thereby contributing to the modulation of neuropathic pain. These findings provide a deeper understanding of the molecular mechanisms underlying neuropathic pain and highlight potential therapeutic targets for its treatment. The differential regulation of this pathway in the spinal cord and dorsal root ganglia (DRG) underscores its complexity and the need for targeted therapeutic strategies.

CircRbms1 fosters MST1 mRNA and protein levels to motivate myocardial ischaemia-reperfusion injury via autophagic status.

AIMS: Acute myocardial infarction (MI) is a significant contributor to death in individuals diagnosed with coronary heart disease on a worldwide level. The specific mechanism by which circRbms1 contributes to the damage caused by myocardial ischaemia-reperfusion (I/R) is not well understood. The primary aim of this study was to examine the role of circRbms1 and its associated mechanisms in the setting of I/R injury. METHODS AND RESULTS: An in vivo MI mice model and an in vitro MI cell model was established. The expression levels were detected using quantitative real-time PCR (qRT-PCR) and western blot. Cellular proliferation, apoptosis, pyroptosis, and autophagy were detected by immunostaining, immunohistochemistry, western blot, and transmission electron microscopy (TEM). Dual-luciferase reporter assay, RNA pull-down assay, and RIP assay were performed to validate the molecular interactions. CircRbms1 was up-regulated in A/R-induced HCMs and acted as a sponge for miR-142-3p, thereby targeting MST1. CircRbms1 could improve stability of MST1 by recruiting IGF2BP2 (all P < 0.05). CircRbms1 knockout reduced cell pyroptosis, improved autophagy and proliferation level in A/R-induced HCMs (all P < 0.05). CircRbms1 knockout alleviated cardiac dysfunction and cell pyroptosis and enhanced autophagy and proliferation in mice through the miR-142-3p/MST1 axis. CONCLUSIONS: CircRbms1 inhibited the miR-142-3p/MST1 axis and played a protective role in myocardial I/R injury. It may provide a new therapeutic target for I/R heart injury.

CircHDAC9 regulates myocardial ischemia-reperfusion injury via miR-671-5p/SOX4 signaling axis.

BACKGROUND: Myocardial ischemia-reperfusion (I/R), a harmful process in the treatment of cardiovascular diseases, can cause secondary damage to the cardiac tissues. Circular RNAs (circRNAs) are important regulators in a number of cardiac disorders. However, the role of circHDAC9 in myocardial I/R injury has not been clarified. METHODS: Human cardiac myocytes (HCMs) were treated with hypoxia/reoxygenation (H/R) and mice were subjected to I/R. Quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR) was used to analyze the expression of circHDAC9, miR-671-5p, and SOX4, and western blot was used to detect SOX4 protein. The binding relationship among circHDAC9, miR-671-5p, and SOX4 was confirmed by RNA pull-down, luciferase, and RNA immunoprecipitation (RIP) assays. The effects of circHDAC9/miR-671-5p/SOX4 axis on the apoptosis, oxidative stress and inflammation were evaluated in both myocardial I/R injury models. RESULTS: The expression of circHDAC9 and SOX4 was noticeably elevated, whereas miR-671-5p expression was downregulated in both myocardial I/R injury models. circHDAC9 knockdown significantly reduced the apoptosis, activities of caspase-3 and caspase-9, ROS intensity, MDA activity, and concentrations of TNF-α, IL-1ß, and IL-6, but increased the viability and SOD activity in H/R-treated HCMs. Suppression of circHDAC9 dramatically reduced the levels of circHDAC9 and SOX4, while enhanced miR-671-5p expression in H/R-treated HCMs. CircHDAC9 functioned via sponging miR-671-5p to regulate SOX4 expression in vitro. Additionally, silencing of circHDAC9 improved the pathological abnormalities and cardiac dysfunction, and reduced the apoptosis, oxidative stress and inflammation in mice with myocardial I/R injury. CONCLUSIONS: Inhibition of circHDAC9 significantly improved myocardial I/R injury by regulating miR-671-5p/SOX4 signaling pathway.

Transcription factor EBF1 mitigates neuropathic pain by rescuing Kv1.2 expression in primary sensory neurons.

Nerve injury-induced alternations of gene expression in primary sensory neurons of the dorsal root ganglion (DRG) are molecular basis of neuropathic pain genesis. Transcription factors regulate gene expression. In this study, we examined whether early B cell factor 1 (EBF1), a transcription factor, in the DRG, participated in neuropathic pain caused by chronic constriction injury (CCI) of the sciatic nerve. EBF1 was distributed exclusively in the neuronal nucleus and coexpressed with cytoplasmic/membrane Kv1.2 in individual DRG neurons. The expression of Ebf1 mRNA and protein was time-dependently downregulated in the ipsilateral lumbar (L) 3/4 DRGs after unilateral CCI. Rescuing this downregulation through microinjection of the adeno-associated virus 5 expressing full-length Ebf1 mRNA into the ipsilateral L3/4 DRGs reversed the CCI-induced decrease of DRG Kv1.2 expression and alleviated the development and maintenance of mechanical, heat and cold hypersensitivities. Conversely, mimicking the downregulation of DRG EBF1 through microinjection of AAV5-expressing Ebf1 shRNA into unilateral L3/4 DRGs produced a reduction of Kv1.2 expression in the ipsilateral L3/4 DRGs, spontaneous pain, and the enhanced responses to mechanical, heat and cold stimuli in naive mice. Mechanistically, EBF1 not only bound to the Kcna2 gene (encoding Kv1.2) promoter but also directly activated its activity. CCI decreased the EBF1 binding to the Kcna2 promoter in the ipsilateral L3/4 DRGs. Our findings suggest that DRG EBF1 downregulation contributes to neuropathic pain likely by losing its binding to Kcna2 promoter and subsequently silencing Kv1.2 expression in primary sensory neurons. Exogenous EBF1 administration may mitigate neuropathic pain by rescuing DRG Kv1.2 expression.

LncRNA RNA ROR Aggravates Hypoxia/Reoxygenation-Induced Cardiomyocyte Ferroptosis by Targeting miR-769-5p/CBX7 Axis.

Ferroptosis is a new way of cell death which is reported to participate in the pathology of myocardial ischemia-reperfusion (MI/R) injury, but it's mechanism remains unclear. The present investigation is to study the emerging role of long non-coding RNA (lncRNA) regulator of reprogramming (ROR) in cardiomyocyte ferroptosis after hypoxia/reoxygenation (H/R) administration. RT-qPCR and/or Western blot methods were performed to examine the gene/or protein levels, and CCK-8, ELISA, and DCFH-DA staining determined the cellular viability and ferroptosis. Dual-luciferase and RNA immunoprecipitation were applied to verify molecular interaction. LncRNA ROR and miR-769-5p were overexpressed and reduced in blood samples from MI patients and H/R-treated AC16 cells, respectively. Mechanistically, lncROR sponged to miR-769-5p, thus upregulating CBX7 expression. Functional experiments presented that lncRNA ROR silence mitigated H/R-stimulated inflammatory damage, oxidative stress, and ferroptosis in AC16 cells, whereas these roles could be reversed by co-downregulation of miR-769-5p or co-overexpression of CBX7. These data uncovered that lncRNA ROR prevented against H/R-induced cardiomyocyte ferroptosis by modulating miR-769-5p/CBX7 signaling, emphasizing the therapeutic value of lncRNA ROR in MI/R injury.

Systemic administration of NIS-lncRNA antisense oligonucleotide alleviates neuropathic pain.

OBJECTIVE: The antisense oligonucleotide (ASO) is an FDA-approved strategy in the treatment of neurological diseases. We have shown the viability of using intrathecal ASO to suppress nerve injury-specific long noncoding RNA (NIS-lncRNA) in dorsal root ganglion (DRG), resulting in a stable and long-lasting antinociceptive effect on NP. This study examined whether systemic administration of NIS-lncRNA ASO relieved the chronic constriction injury (CCI)-induced nociceptive hypersensitivity. METHODS: A single subcutaneous injection of NIS-lncRNA ASO at a dose of 1,000 µg was carried out 7 days after CCI or sham surgery in male mice. Behavioral tests were performed one day before surgery and at different days after surgery. DRG and spinal cord were finally collected for quantitative real-time RT-PCR and Western blot assays. RESULTS: NIS-lncRNA ASO significantly alleviated CCI-induced mechanical allodynia, heat hyperalgesia, and cold hyperalgesia starting on day 14 or 21 post-ASO injection and lasting for at least 7 days on the ipsilateral side. Additionally, CCI-induced spontaneous pain and ipsilateral dorsal horn neuronal and astrocyte hyperactivation were blocked on day 28 after NIS-lncRNA ASO injection. As predicted, the CCI-induced increases in the levels of NIS-lncRNA and its downstream target C-C motif chemokine ligand 2 in the ipsilateral lumbar 3 and 4 DRGs were attenuated on day 28 following NIS-lncRNA ASO injection. CONCLUSION: Our findings indicate that systemic administration of NIS-lncRNA ASO also produces a stable and long-lasting antinociceptive effect on neuropathic pain. NIS-lncRNA ASO may have potential clinical application in the treatment of this disorder.

Sensory neuron-specific long noncoding RNA in small non-peptidergic dorsal root ganglion neurons selectively impairs nerve injury-induced mechanical hypersensitivity.

AIMS: Nerve injury-induced mechanical hypersensitivity is one of major clinical symptoms in neuropathic pain patients. Understanding molecular mechanisms underlying this symptom is crucial for developing effective therapies. The present study was to investigate whether sensory neuron-specific long noncoding RNA (SS-lncRNA) predominantly expressed in small non-peptidergic dorsal root ganglion (DRG) neurons repaired nerve injury-induced mechanical hypersensitivity. MATERIALS AND METHODS: SS-lncRNA downregulation in the mas-related G protein-coupled receptor member D (Mrgprd)-expressed DRG neurons was rescued and mimicked by crossbreeding MrgprdCreERT2/+ lines with Rosa26SS-lncRNA knock-in mice and SS-lncRNAfl/fl mice, respectively, followed by tamoxifen injection. KEY FINDINGS: Rescuing SS-lncRNA downregulation in the Mrgprd-expressed DRG neurons significantly reversed the spinal nerve ligation (SNL)-induced reduction of the calcium-activated potassium channel subfamily N member 1 (KCNN1) in these DRG neurons and alleviated the SNL-induced mechanical hypersensitivity, without affecting the SNL-induced heat and cold nociceptive hypersensitivities, on the ipsilateral side. Conversely, mimicking SS-lncRNA downregulation in the Mrgprd-expressed DRG neurons reduced basal KCNN1 expression in these DRG neurons and produced the enhanced response to mechanical stimulation, but not thermal and cold stimuli, on bilateral sides. Mechanistically, SS-lncRNA downregulation caused a reduction in its binding to lysine-specific demethylase 6B (KDM6B) and consequent recruitment of less KDM6B to Kcnn1 promoter and an increase of H3K27me3 enrichment in this promoter in injured DRG. SIGNIFICANCE: Our findings suggest that SS-lncRNA downregulation in small non-peptidergic sensory neurons is required specifically for nerve injury-induced mechanical hypersensitivity likely through silencing KCNN1 expression caused by KDM6B-gated increase of H3K27me3 enrichment in Kcnn1 promoter in these neurons.

A sensory neuron-specific long non-coding RNA reduces neuropathic pain by rescuing KCNN1 expression.

Nerve injury to peripheral somatosensory system causes refractory neuropathic pain. Maladaptive changes of gene expression in primary sensory neurons are considered molecular basis of this disorder. Long non-coding RNAs (lncRNAs) are key regulators of gene transcription; however, their significance in neuropathic pain remains largely elusive.Here, we reported a novel lncRNA, named sensory neuron-specific lncRNA (SS-lncRNA), for its expression exclusively in dorsal root ganglion (DRG) and trigeminal ganglion. SS-lncRNA was predominantly expressed in small DRG neurons and significantly downregulated due to a reduction of early B cell transcription factor 1 in injured DRG after nerve injury. Rescuing this downregulation reversed a decrease of the calcium-activated potassium channel subfamily N member 1 (KCNN1) in injured DRG and alleviated nerve injury-induced nociceptive hypersensitivity. Conversely, DRG downregulation of SS-lncRNA reduced the expression of KCNN1, decreased total potassium currents and afterhyperpolarization currents and increased excitability in DRG neurons and produced neuropathic pain symptoms.Mechanistically, downregulated SS-lncRNA resulted in the reductions of its binding to Kcnn1 promoter and heterogeneous nuclear ribonucleoprotein M (hnRNPM), consequent recruitment of less hnRNPM to the Kcnn1 promoter and silence of Kcnn1 gene transcription in injured DRG.These findings indicate that SS-lncRNA may relieve neuropathic pain through hnRNPM-mediated KCNN1 rescue in injured DRG and offer a novel therapeutic strategy specific for this disorder.

Silencing RIPK1/mTORC1 signalling attenuated the inflammation and oxidative stress in diabetic cardiomyopathy.

BACKGROUND: Diabetes cardiomyopathy (DCM) is one of the major risk factors for the heart failure of the diabetic patients. RIPK1 maybe participate in the regulation of the oxidative stress and inflammation during DCM. METHODS: H&E and Masson staining were utilized to assess the inflammation and fibrosis in myocardial tissues. CCK-8 and TUNEL staining were utilized to analyze cell viability and apoptosis, respectively. SOD activity and MDA content were detected utilizing the kits. The formation of autophagosomes was measured by immunofluorescence assay. RESULTS: RIPK1 and RPTOR (a component of mTORC1) expression and oxidative stress level were upregulated, but autophagy was decreased in the myocardial tissues of DCM rat characterized by the high body weight and blood glucose, abnormal cardiac function, myocardial inflammation and fibrosis. High glucose (HG) treatment resulted in cell viability and autophagy level decreasing, inflammatory cytokines expression increasing and oxidative stress increasing in cardiac fibroblasts (CFs). Meanwhile, RIPK1 and RPTOR expression also was increased in HG-treated cells. HG-induced CFs apoptosis, inflammation, oxidative stress and the inhibition of HG to cell viability and autophagy was partly reversed by the inhibitor of RIPK1 and mTORC1. CONCLUSION: Overall, RIPK1/mTORC1 signalling suppression improved HG-induced apoptosis, inflammation and oxidative stress through activation autophagy. These data provided a reliable evidence that RIPK1 may be a potential target for DCM therapeutic.

Knockout of circRNA single stranded interacting protein 1 (circRBMS1) played a protective role in myocardial ischemia-reperfusion injury though inhibition of miR-2355-3p/Mammalian Sterile20-like kinase 1 (MST1) axis.

Evidence suggests circRBMS1 regulates mRNA to mediate cell apoptosis, inflammation, and oxidative stress in different diseases. MST1 is reported to be the target and activator of apoptosis-related molecules and signaling pathways. Hence, the present study aims to investigate the role of circ-RBMS1/miR-2355-3p/MST1 in the development of I/R injury. In vitro experiments showed increased circ-RBMS1 and decreased miR-2355-3p in H/R-induced HCMs. CircRBMS1 served as a sponge for miR-2355-3p and miR-2355-3p targeted MST1. Furthermore, knockout of circRBMS1 attenuated cell apoptosis, oxidized stress, and inflammation in H/R-induced HCMs. In vivo experiments indicated circRBMS1 knockdown attenuated cardiac function damage, cell apoptosis, oxidative stress injury and inflammatory response through miR-2355-3p/MST1 axis in mice. In summary, these results demonstrated circRBMS1 played a protective role in myocardial I/R injury though inhibition of miR-2355-3p/MST1 axis. It might provide a new therapeutic target for cardiac I/R injury.

The effectiveness of repetitive paravertebral block with ropivacaine and dexmedetomidine for the prevention of postherpetic neuralgia in patients with acute herpes zoster.

Introduction: Herpes zoster (HZ) is a disease caused by the reactivation of the varicella zoster virus. Postherpetic neuralgia (PHN) is the most common complication of HZ. Aim: Repetitive paravertebral block with local anaesthetics and dexmedetomidine for the prevention of PHN in patients with acute herpes zoster. Material and methods: 104 patients with acute herpes zoster were randomly divided into two groups. Group Rop received repetitive paravertebral block with 0.25% ropivacaine 20 ml per 72 h three times. Group Dex received repetitive paravertebral block with a mixture of 0.25% ropivacaine 20 ml and dexmedetomidine 20 µg per 72 h three times. Patients were permitted to take tramadol when the visual analogue scale (VAS) ≥ 4. The incidence of zoster-related pain was recorded at 1, 3, and 6 months after the end of treatments; VAS scores and the dose of rescue drug were recorded at 1 week, 2 weeks, 1 month, 3 months, and 6 months after the end of treatments. Results: At 1 month post therapy, the incidence of zoster-related pain was 11% in Group Dex, compared with 35% in Group Rop (p = 0.005). At 3 months post therapy, the incidence of zoster-related pain in Group Dex was still significantly lower than in Group Rop. The VAS scores and the dose of rescue drug in Group Dex were also significantly lower than in Group Rop at each time point (p < 0.05). Conclusions: Repetitive paravertebral block with local anaesthetics and dexmedetomidine in patients with acute herpes zoster can significantly reduce the incidence of zoster-related pain.

LncRNA ROR modulates myocardial ischemia-reperfusion injury mediated by the miR-185-5p/CDK6 axis.

LncRNAs and miRNAs are correlated with the pathogenesis of myocardial ischemia-reperfusion injury (MIRI). Whether lncRNA ROR or miR-185-5p plays a crucial role in MIRI is still unclear. In in-vitro, human cardiac myocytes (HCMs) were treated with hypoxia/reoxygenation (H/R). Wistar rats were used to set up an in-vitro I/R model by means of recanalization after ligation. Evaluation of the myocardial injury marker lactate dehydrogenase (LDH) in HCMs cells was performed. The expression of miR-185-5p and ROR, IL-1ß, and IL-18 were detected by qRT-PCR. ELISA was also performed to evaluate the secretion of IL-1ß and IL-18. Western blotting was carried out to determine CDK6, NLRP3, GSDMD-N, ASC, and cleaved-caspase1 protein expression. The relationship between miR-185-5p and CDK6 or ROR was confirmed by a dual-luciferase reporter assay. Our findings revealed that H/R treated HCMs showed a significantly decreased miR-185-5p expression and increased expression of CDK6 and ROR. ROR knockdown reduced H/R induced pyroptosis and inflammation, while knockdown of miR-185-5p accelerated the effect. Furthermore, miR-185-5p was negatively regulated and absorbed by ROR in HCMs. Overexpression of miR-185-5p reversed the H/R-induced cell pyroptosis and upregulation of LDH, IL-1ß, and IL-18. In HCMs, miR-185-5p was also negatively regulated and related to CDK6 expression. Moreover, overexpression of CDK6 significantly inhibited the effects of miR-185-5p mimics on the inflammatory response and pyroptosis of HCMs. Knockdown of ROR alleviated H/R-induced myocardial injury by elevating miR-185-5p and inhibiting CDK6 expression. Taken together, our results show that the ROR/miR-185-5p/CDK6 axis modulates cell pyroptosis induced by H/R and the inflammatory response of HCMs.

Long non-coding RNA ROR sponges miR-138 to aggravate hypoxia/reoxygenation-induced cardiomyocyte apoptosis via upregulating Mst1.

BACKGROUND: Hypoxia/reoxygenation (H/R) injury of cardiomyocytes causes an irreversible damage to heart and largely results in acute myocardial infarction. Study has indicated lncRNA ROR aggravates myocardial ischemia/reperfusion (I/R) injury. Also, lncRNA ROR sponges miR-138 to promote osteogenesis. MiR-138 involves in hypoxic pulmonary vascular remodelling by targeting Mst1. However, the interaction between lncRNA ROR, miR-138 and Mst1 involved in myocardial H/R injury is still unknown. METHODS: H9C2 cells were used to establish H/R injury model. The expression levels of lncRNA ROR and miR-138 were modified by transfection with the miR-138 mimics or lncRNA ROR overexpression plasmid. MTT and flow cytometry analysis were performed to detect cell proliferation and apoptosis. Dual luciferase reporter assay was used to determine interaction between lncRNA ROR and miR-138 or miR-138 and Mst1. Expression levels of lncRNA ROR, miR-138, Mst1 and apoptosis-related markers were determined by qRT-PCR or western blotting. RESULTS: LncRNA ROR was significantly up-regulated, while miR-138 was obviously down-regulated in H/R-induced injury of H9C2 cells. Furthermore, miR-138 overexpression alleviated cardiac cell apoptosis induced by H/R injury. Mst1 was revealed to be a target of miR-138 and negatively regulated by miR-138. Mst1 overexpression reversed the protective effects of miR-138 on H/R injury of H9C2 cells. LncRNA ROR was identified as a sponge for miR-138. MiR-138 could protect H9C2 cells form H/R injury induced by lncRNA ROR overexpression. CONCLUSION: Our study provides that lncRNA ROR sponges miR-138 to aggravate H/R-induced myocardial cell injury by upregulating the expression of Mst1.

The lncRNA ROR/miR-124-3p/TRAF6 axis regulated the ischaemia reperfusion injury-induced inflammatory response in human cardiac myocytes.

Myocardial ischaemia reperfusion injury (MIRI) is considered the primary cause of death in patients with cardiovascular diseases. Recently, long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) have been found to be involved in the pathogenesis of MIRI. However, whether lncRNA ROR and miR-124-3p play roles in MIRI and the underlying mechanism remain undetermined. HCMs were exposed to hypoxic conditions for 2 h followed by re-oxygenation (H/R) treatment. Expression of miR-124-3p and lncRNA ROR in HCMs was measured by qRT-PCR. TRAF6 expression was evaluated by qRT-PCR and western blotting. ELISA and qRT-PCR were conducted to assess the production of TNF-α, IL-6, and IL-1ß. The interaction between miR-124-3p and TRAF6, as well as between miR-124-3p and lncRNA ROR, was verified by dual-luciferase reporter assay. Cell apoptosis was detected by flow cytometry analysis. Our data revealed that miR-124-3p was significantly downregulated, while TRAF6 and lncRNA ROR were upregulated in both MIRI rat model and H/R treated HCMs. Overexpression of miR-124-3p reversed the H/R-induced cell apoptosis and upregulation of TNF-α, IL-6, and IL-1ß. Mechanistically, miR-124-3p bound and negatively regulated TRAF6 expression in HCMs. Moreover, TRAF6 overexpression significantly blocked the effects of miR-124-3p mimics on cell apoptosis and inflammatory response of HCMs, which involved the NF-κB pathway. Further analysis showed that lncRNA ROR sponged and negatively regulated miR-124-3p in HCMs. Overexpression of IL-1ß was demonstrated to promote H/R induced cell apoptosis in HCMs. In addition, overexpression of ROR further enhanced the H/R-induced inflammation and cell apoptosis through its action on miR-124-3p. The lncRNA ROR/miR-124-3p/TRAF6 axis regulated the H/R-induced cell apoptosis and inflammatory response of HCMs.

Chlorogenic-induced inhibition of non-small cancer cells occurs through regulation of histone deacetylase 6.

Chlorogenic acid (CGA), an ester with various pharmacological effects, is important in cancer therapy. However, the specific antitumor mechanism of CGA is not entirely clear, especially with respect to its suppression of non-small cell lung cancer (NSCLC). The present study was carried out to assess the effect of CGA on NSCLC, and the mechanism involved. Cell viability assay and colony formation assay revealed that CGA blocked the proliferative capacity of NSCLC cells in vitro. Results from the migration assay suggested that CGA also inhibited the migration of A549 cells. Other assays further revealed that CGA strongly and selectively inhibited histone deacetylase 6 (HDAC6) activity and suppressed the activity of matrix metalloproteinase-2 (MMP-2) through decreased expression of Ac-NF-κB. Tumorigenicity assay showed that CGA also inhibited the proliferation and metabolism of NSCLC in vivo. These results indicate that CGA significantly suppresses the proliferation of NSCLC by regulating the activity of histone deacetylase 6.

Naringenin pre-treatment inhibits neuroapoptosis and ameliorates cognitive impairment in rats exposed to isoflurane anesthesia by regulating the PI3/Akt/PTEN signalling pathway and suppressing NF-κB-mediated inflammation.

The volatile anaesthetic isoflurane is one of the most frequently employed general anaesthetics in neonates, children and adults. Accumulating evidence demonstrated that exposure to anaesthetics is associated with widespread neurodegeneration and cognitive impairment. Thus, the identification and development of compounds capable of preventing or reducing these adverse effects is of great clinical importance. For this purpose, the present study aimed to assess the effects of a flavonoid, naringenin, on isoflurane-induced neuroapoptosis and cognitive impairment. Separate groups of neonatal rat pups were administered naringenin at 25, 50 or 100 mg/kg body weight from postnatal day 1 (P1) to P21. On P7, the pups were exposed to 6 h of isoflurane (0.75%) anaesthesia. Neuroapoptosis was examined using the TUNEL assay. The expression of cleaved caspase-3, the apoptotic pathway proteins (Bad, Bax, Bcl-2 and Bcl-xL), the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) pathway proteins [Akt, phosphorylated (-)Akt, glycogen synthase kinase 3ß (GSK­3ß), p­GSK-3ß, phosphatase and tensin homolog (PTEN)] and nuclear factor-κB (NF-κB)­mediated signalling proteins were determined by western blot analysis. General behaviour, as well as the learning ability and memory of the pups were assessed. Naringenin significantly inhibited isoflurane­induced neuroapoptosis and markedly decreased the protein expression of caspase-3, Bad, Bax, NF-κB, tumor necrosis factor-α, interleukin (IL)-6 and IL-1ß. Furthermore, naringenin increased the expression of Bcl-xL and Bcl-2 and activated the PI3K/Akt pathway. Significant improvements in learning capacity and memory retention were observed following naringenin treatment. Naringenin effectively ameliorated cognitive dysfunction and reduced isoflurane­induced apoptosis as well as modulating the PI3/Akt/PTEN and NF-κB signalling pathways.

Phosphorylated neuronal nitric oxide synthase in neuropathic pain in rats.

Neuropathic pain caused by nervous system damage or system dysfunction. The pathogenesis and the mechanism underlying neuropathic pain remains unclear. The only known neurobiological component involved in the neuropathic pain is nitric oxide (NO). NO is synthesized by nitric oxide synthase (nNOS) from L-arginine and oxygen. nNOS is involved in the inflammatory pain and neuropathic pain. In this study, we aimed to identify whether KN93 reduced the pain in the rats. Sixty adult male SD rat were randomly divided into 4 groups. Sham group and model group were not received treatment. Experimental group received intrathecal injection of KN93, and negative control group received DMSO injection 30 min before pain test. After last test of pain threshold, the rats were sacrificed and lumbar spinal tissues were sampled for analysis of the expression of pnNOS and pCaMK II by quantitative PCR and Western blotting. Pain threshold was increased in the rats received KN93 treatment (P<0.01), and the expression levels of pnNOS was increased (P<0.05) in experimental group and accompanied with decrease of CaMK II expression (P<0.05). By administration of KN93, the interaction of nNOS and the adaptor protein CAPON was reduced through inhibition of CaMK II by KN93. In conclusion, this study reveals that KN93 can reduce neuropathic pain via inhibiting the activity of CaMK II, and then increase the level of phosphorylated nNOS, to reduce the interaction with CAPON.

[Activity of four cry gene promoters in spoIIID mutant of Bacillus thuringiensis].

OBJECTIVE: We studied the influence of spoIIID gene deletion on the activity of cry1Ac, cry3A, cry4A and cry8E gene promoters in Bacillus thuringiensis and compared the activity among these promoters in spoIIID mutant (HD-delta SpoIIID). METHODS: We constructed 4 promoter fusions with lacZ gene and transformed them into wild-type strain HD-73 and HD-deltaSpoIIID to analyze their transcriptional activity. We constructed a spoIIID gene mutant (HD-(deltaASpoIIID with deletion of the cry1Ac-harboring native plasmid based on HD-ASpoIIID strain. We constructed four promoter fusions with crylAc gene and transformed them into HD-ASpoIIID and HD(-)-deltaSpoIIID to perform Cry protein quantization and bioassay. RESULTS: By Beta-galactosidase assay we found that the activities of the four promoters were, in decreasing order, Pcry8E > Pcry1A > Pcry4A > Pcry3A in both HD-73 and HD-deltaSpoIIID strains. The deletion of spoIID had no effect on transcriptional activity of PcrylAc and Pcry8E. The transcriptional activity of Pcry3A in HD-deltaSpoIIID was slightly higher than that in HD-73. The transcriptional activity of Pcry4A in HD-deltaSpoIIID was decreased compared to HD-73. The Cry1Ac protein production directed by Pcry1Ac was as much as Pcry8E in HD-deltaSpoIIID and higher than that by Pcry4A and Pcry3A in accordance with the bioassay result. CONCLUSION: The cry8E gene promoter is the strongest promoter among four promoters in spoIIID gene mutant at transcriptional level. The Cry1 Ac protein production directed by Pcry1Ac is almost equal to that by Pcry8E.