Genetic and phenotypic profiling of single living circulating tumor cells from patients with microfluidics.

Accurate prediction of the efficacy of immunotherapy for cancer patients through the characterization of both genetic and phenotypic heterogeneity in individual patient cells holds great promise in informing targeted treatments, and ultimately in improving care pathways and clinical outcomes. Here, we describe the nanoplatform for interrogating living cell host-gene and (micro-)environment (NICHE) relationships, that integrates micro- and nanofluidics to enable highly efficient capture of circulating tumor cells (CTCs) from blood samples. The platform uses a unique nanopore-enhanced electrodelivery system that efficiently and rapidly integrates stable multichannel fluorescence probes into living CTCs for in situ quantification of target gene expression, while on-chip coculturing of CTCs with immune cells allows for the real-time correlative quantification of their phenotypic heterogeneities in response to immune checkpoint inhibitors (ICI). The NICHE microfluidic device provides a unique ability to perform both gene expression and phenotypic analysis on the same single cells in situ, allowing us to generate a predictive index for screening patients who could benefit from ICI. This index, which simultaneously integrates the heterogeneity of single cellular responses for both gene expression and phenotype, was validated by clinically tracing 80 non-small cell lung cancer patients, demonstrating significantly higher AUC (area under the curve) (0.906) than current clinical reference for immunotherapy prediction.

Structural basis for the allosteric regulation and dynamic assembly of DNMT3B.

Oligomerization of DNMT3B, a mammalian de novo DNA methyltransferase, critically regulates its chromatin targeting and DNA methylation activities. However, how the N-terminal PWWP and ADD domains interplay with the C-terminal methyltransferase (MTase) domain in regulating the dynamic assembly of DNMT3B remains unclear. Here, we report the cryo-EM structure of DNMT3B under various oligomerization states. The ADD domain of DNMT3B interacts with the MTase domain to form an autoinhibitory conformation, resembling the previously observed DNMT3A autoinhibition. Our combined structural and biochemical study further identifies a role for the PWWP domain and its associated ICF mutation in the allosteric regulation of DNMT3B tetramer, and a differential functional impact on DNMT3B by potential ADD-H3K4me0 and PWWP-H3K36me3 bindings. In addition, our comparative structural analysis reveals a coupling between DNMT3B oligomerization and folding of its substrate-binding sites. Together, this study provides mechanistic insights into the allosteric regulation and dynamic assembly of DNMT3B.

DNA conformational dynamics in the context-dependent non-CG CHH methylation by plant methyltransferase DRM2.

DNA methylation provides an important epigenetic mechanism that critically regulates gene expression, genome imprinting, and retrotransposon silencing. In plants, DNA methylation is prevalent not only in a CG dinucleotide context but also in non-CG contexts, namely CHG and CHH (H = C, T, or A) methylation. It has been established that plant non-CG DNA methylation is highly context dependent, with the +1- and +2-flanking sequences enriched with A/T nucleotides. How DNA sequence, conformation, and dynamics influence non-CG methylation remains elusive. Here, we report structural and biochemical characterizations of the intrinsic substrate preference of DOMAINS REARRANGED METHYLTRANSFERASE 2 (DRM2), a plant DNA methyltransferase responsible for establishing all cytosine methylation and maintaining CHH methylation. Among nine CHH motifs, the DRM2 methyltransferase (MTase) domain shows marked substrate preference toward CWW (W = A or T) motifs, correlating well with their relative abundance in planta. Furthermore, we report the crystal structure of DRM2 MTase in complex with a DNA duplex containing a flexible TpA base step at the +1/+2-flanking sites of the target nucleotide. Comparative structural analysis of the DRM2-DNA complexes provides a mechanism by which flanking nucleotide composition impacts DRM2-mediated DNA methylation. Furthermore, the flexibility of the TpA step gives rise to two alternative DNA conformations, resulting in different interactions with DRM2 and consequently temperature-dependent shift of the substrate preference of DRM2. Together, this study provides insights into how the interplay between the conformational dynamics of DNA and temperature as an environmental factor contributes to the context-dependent CHH methylation by DRM2.

Ternary Heteroatomic Doping Induced Microenvironment Engineering of Low Fe-N4-Loaded Carbon Nanofibers for Bifunctional Oxygen Electrocatalysis.

Fabricating highly efficient and long-life redox bifunctional electrocatalysts is vital for oxygen-related renewable energy devices. To boost the bifunctional catalytic activity of Fe-N-C single-atom catalysts, it is imperative to fine-tune the coordination microenvironment of the Fe sites to optimize the adsorption/desorption energies of intermediates during oxygen reduction/evolution reactions (ORR/OER) and simultaneously avoid the aggregation of atomically dispersed metal sites. Herein, a strategy is developed for fabricating a free-standing electrocatalyst with atomically dispersed Fe sites (≈0.89 wt.%) supported on N, F, and S ternary-doped hollow carbon nanofibers (FeN4 -NFS-CNF). Both experimental and theoretical findings suggest that the incorporation of ternary heteroatoms modifies the charge distribution of Fe active centers and enhances defect density, thereby optimizing the bifunctional catalytic activities. The efficient regulation isolated Fe centers come from the dual confinement of zeolitic imidazole framework-8 (ZIF-8) and polymerized ionic liquid (PIL), while the precise formation of distinct hierarchical three-dimensional porous structure maximizes the exposure of low-doping Fe active sites and enriched heteroatoms. FeN4 -NFS-CNF achieves remarkable electrocatalytic activity with a high ORR half-wave potential (0.90 V) and a low OER overpotential (270 mV) in alkaline electrolyte, revealing the benefit of optimizing the microenvironment of low-doping iron single atoms in directing bifunctional catalytic activity.

Electromagnetic field and cardiovascular diseases: A state-of-the-art review of diagnostic, therapeutic, and predictive values.

Despite encouraging advances in early diagnosis and treatment, cardiovascular diseases (CVDs) remained a leading cause of morbidity and mortality worldwide. Increasing evidence has shown that the electromagnetic field (EMF) influences many biological processes, which has attracted much attention for its potential therapeutic and diagnostic modalities in multiple diseases, such as musculoskeletal disorders and neurodegenerative diseases. Nonionizing EMF has been studied as a therapeutic or diagnostic tool in CVDs. In this review, we summarize the current literature ranging from in vitro to clinical studies focusing on the therapeutic potential (external EMF) and diagnostic potential (internal EMF generated from the heart) of EMF in CVDs. First, we provided an overview of the therapeutic potential of EMF and associated mechanisms in the context of CVDs, including cardiac arrhythmia, myocardial ischemia, atherosclerosis, and hypertension. Furthermore, we investigated the diagnostic and predictive value of magnetocardiography in CVDs. Finally, we discussed the critical steps necessary to translate this promising approach into clinical practice.

Nivolumab combined docetaxel versus nivolumab in patients with previously treated nonsmall cell lung cancer: a phase 2 study.

The current standard second-line treatment is immune checkpoint inhibitors monotherapy for nonsmall cell lung cancer (NSCLC) patients. The objective of this phase 2 study was to evaluate the efficacy and safety of nivolumab plus docetaxel compared with nivolumab monotherapy for second-line therapy in immunotherapy-naive patients with advanced NSCLC. Progression-free survival (PFS) was the primary endpoint of this phase 2 study. Patients were randomized to receive nivolumab plus docetaxel or nivolumab monotherapy. From July 2019 to June 2022, a total of 22 patients were recruited, with significantly longer median PFS observed in the nivolumab plus docetaxel group (4.0 months) compared to the nivolumab group (2.0 months), P  = 0.0019. The study was closed in June 2022 due to slow recruitment. The objective response rate was 10.0% [95% confidence interval (CI), 0-28.6] in the nivolumab group and 25% (95% CI, 0.5-49.5) in the nivolumab + docetaxel group ( P  = 0.346). Disease control was significantly higher in the nivolumab plus docetaxel arm (40.0% versus 83.3%, P  = 0.035). There was also an improvement in overall survival (OS) in the nivolumab + docetaxel arm, but this was not statistically significant (10.0 months versus 7.2 months, P  = 0.129). The addition of docetaxel to nivolumab was well-tolerated, with adverse events more common in the combination group. Despite the small sample size, the results suggest that the addition of docetaxel to nivolumab may be a promising treatment option for NSCLC patients progressing on platinum-based chemotherapy, with trends towards improved OS observed.

Tuning d-orbitals to control spin-orbit coupling in terminated MXenes.

Theory and experiment have revealed that spin-orbit coupling (SOC) strongly depends on the relativistic effect in topological insulators (TIs), while the influence of orbitals is always ignored. Herein, we provide a direct way of controlling effective SOC with the help of orbital effects, reducing the dependence on elements. Taking 5d W2CO2 and 4d Mo2CO2 MXenes as a specific example, we predict that by decreasing the hybridization strength of W atoms with C or O atoms in 2D W2CO2, the nontrivial bandgaps at the Γ-point are directly enhanced. The weak hybridization of W atoms with ligand elements enhances the electron localization of degenerate d-orbitals of three groups under the triangular prism crystal field, inducing stronger on-site Coulomb repulsion that enhances orbital polarization as well as boosts the SOC effect. Meanwhile, similar results have also been observed in 4d Mo2CO2. This implies that the orbital effects are an efficient and straightforward way to control the nontrivial bandgap in 2D MXene TIs. Our work not only provides an alternative perspective on designing large nontrivial bandgaps but also brings a possibility to control the SOC effect for TI devices.

LINC00624 affects hepatocellular carcinoma proliferation and apoptosis through the miR-342-3p/DNAJC5 axis.

LINC00624 is a long noncoding RNA (lncRNA) which was seldom investigated before. The goal of our study is to clarify the expression and underlying network of LINC00624 in hepatocellular carcinoma (HCC). Here, both HCC and normal living cell lines were employed. Real-time quantitative PCR and western blot were used to determine the pattern of genes and proteins. Colony formation, flow cytometry and western blot tests were used to determine cell proliferation and apoptosis, respectively. Dual luciferase was used to verify molecule-molecule interactions. LINC00624 expression was increased in HCC cell lines and miR-342-3p was decreased. Elimination of LINC00624 increased proliferation while decreasing cell apoptosis. LINC00624 acted as a molecular sponge for miR-342-3p, hence facilitating DNAJC5 expression. Functional tests demonstrated that miR-342-3p suppression could reverse the effect of LINC00624 silence and overexpression of DNAJC5 significantly mitigated the biological consequences of miR-342-3p. These finding demonstrated that LINC00624 aggravated HCC progression by modulating proliferation and apoptosis via targeting miR-342-3p/DNAJC5 axis. These data support that inhibition of LINC00624 may a potential treatment strategies of HCC.

Improvements to a GLCM-based machine-learning approach for quantifying posterior capsule opacification.

BACKGROUND: Posterior capsular opacification (PCO) is a common complication following cataract surgery that leads to visual disturbances and decreased quality of vision. The aim of our study was to employ a machine-learning methodology to characterize and validate enhancements applied to the grey-level co-occurrence matrix (GLCM) while assessing its validity in comparison to clinical evaluations for evaluating PCO. METHODS: One hundred patients diagnosed with age-related cataracts who were scheduled for phacoemulsification surgery were included in the study. Following mydriasis, anterior segment photographs were captured using a high-resolution photographic system. The GLCM was utilized as the feature extractor, and a supported vector machine as the regressor. Three variations, namely, GLCM, GLCM+C (+axial information), and GLCM+V (+regional voting), were analyzed. The reference value for regression was determined by averaging clinical scores obtained through subjective analysis. The relationships between the predicted PCO outcome scores and the ground truth were assessed using Pearson correlation analysis and a Bland-Altman plot, while agreement between them was assessed through the Bland-Altman plot. RESULTS: Relative to the ground truth, the GLCM, GLCM+C, and GLCM+V methods exhibited correlation coefficients of 0.706, 0.768, and 0.829, respectively. The relationship between the PCO score predicted by the GLCM+V method and the ground truth was statistically significant (p < 0.001). Furthermore, the GLCM+V method demonstrated competitive performance comparable to that of two experienced clinicians (r = 0.825, 0.843) and superior to that of two junior clinicians (r = 0.786, 0.756). Notably, a high level of agreement was observed between predictions and the ground truth, without significant evidence of proportional bias (p > 0.05). CONCLUSIONS: Overall, our findings suggest that a machine-learning approach incorporating the GLCM, specifically the GLCM+V method, holds promise as an objective and reliable tool for assessing PCO progression. Further studies in larger patient cohorts are warranted to validate these findings and explore their potential clinical applications.

IMpower210: A phase III study of second-line atezolizumab vs. docetaxel in East Asian patients with non-small cell lung cancer.

Objective: IMpower210 (NCT02813785) explored the efficacy and safety of single-agent atezolizumab vs. docetaxel as second-line treatment for advanced non-small cell lung cancer (NSCLC) in East Asian patients. Methods: Key eligibility criteria for this phase III, open-label, randomized study included age ≥18 years; histologically documented advanced NSCLC per the Union for International Cancer Control/American Joint Committee on Cancer staging system (7th edition); Eastern Cooperative Oncology Group performance status of 0 or 1; and disease progression following platinum-based chemotherapy for advanced or metastatic NSCLC. Patients were randomized 2:1 to receive either atezolizumab (1,200 mg) or docetaxel (75 mg/m2). The primary study endpoint was overall survival (OS) in the intention-to-treat (ITT) population with wild-type epidermal growth factor receptor expression (ITT EGFR-WT) and in the overall ITT population. Results: Median OS in the ITT EGFR-WT population (n=467) was 12.3 [95% confidence interval (95% CI), 10.3-13.8] months in the atezolizumab arm (n=312) and 9.9 (95% CI, 7.8-13.9) months in the docetaxel arm [n=155; stratified hazard ratio (HR), 0.82; 95% CI, 0.66-1.03]. Median OS in the overall ITT population was 12.5 (95% CI, 10.8-13.8) months with atezolizumab treatment and 11.1 (95% CI, 8.4-14.2) months (n=377) with docetaxel treatment (n=188; stratified HR, 0.87; 95% CI, 0.71-1.08). Grade 3/4 treatment-related adverse events (TRAEs) occurred in 18.4% of patients in the atezolizumab arm and 50.0% of patients in the docetaxel arm. Conclusions: IMpower210 did not meet its primary efficacy endpoint of OS in the ITT EGFR-WT or overall ITT populations. Atezolizumab was comparatively more tolerable than docetaxel, with a lower incidence of grade 3/4 TRAEs.

Merging of Light/Dark Palladium Catalytic Cycles Enables Multicomponent Tandem Alkyl Heck/Tsuji-Trost Homologative Amination Reaction toward Allylic Amines.

A visible light-induced palladium-catalyzed homologative three-component synthesis of allylic amines has been developed. This protocol proceeds via a unique mechanism involving two distinct cycles enabled by the same Pd(0) catalyst: a visible light-induced hybrid radical alkyl Heck reaction between 1,1-dielectrophile and styrene, followed by the "in dark" classical Tsuji-Trost-type allylic substitution reaction. This method works well with a broad range of primary and secondary amines, aryl alkenes, dielectrophiles, and in complex settings. The regiochemistry of the obtained products is primarily governed by the structure of 1,1-dielectrophile. Involvement of π-allyl palladium intermediates allowed for the control of stereoselectivity, which has been demonstrated with up to 95:5 er.

Results of the phase IIa study to evaluate the efficacy and safety of rezivertinib (BPI-7711) for the first-line treatment of locally advanced or metastatic/recurrent NSCLC patients with EGFR mutation from a phase I/IIa study.

BACKGROUND: Rezivertinib (BPI-7711) is a novel third-generation epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI). This phase IIa study was part of a phase I/IIa study (NCT03386955), aimed to evaluate the efficacy and safety of rezivertinib as the first-line treatment for patients with locally advanced or metastatic/recurrent EGFR mutated non-small cell lung cancer (NSCLC). METHODS: Patients received the first-line treatment of 180 mg rezivertinib orally once daily until disease progression, unacceptable toxicity, or withdrawal of consent. The primary endpoint was the objective response rate (ORR) assessed by blinded independent central review (BICR). Secondary endpoints included disease control rate (DCR), duration of response (DoR), progression-free survival (PFS), overall survival (OS), and safety. RESULTS: From Jun 12, 2019, to Oct 17, 2019, 43 patients were enrolled. At the data cutoff date on Dec 23, 2021, the ORR by BICR was 83.7% (95% CI: 69.3-93.2%). The median DoR was 19.3 (95% CI: 15.8-25.0) months. The median PFS by BICR was 20.7 (95% CI: 13.8-24.8) months and 22.0 (95% CI: 16.8-26.3) months by investigators. Data on OS was immature. Totally, 40 (93.0%) patients had at least one treatment-related adverse event while 4 (9.3%) of them were grade ≥ 3. CONCLUSIONS: Rezivertinib (BPI-7711) showed promising efficacy and a favorable safety profile for the treatment among the locally advanced or metastatic/recurrent NSCLC patients with EGFR mutation in the first-line setting. TRIAL REGISTRATION: ClinicalTrials.gov, NCT03386955.

Synergistic Solar-Driven Freshwater Generation and Electricity Output Empowered by Wafer-Scale Nanostructured Silicon.

Electricity generation triggered by the ubiquitous water evaporation process provides an intriguing way to harvest energy from water. Meanwhile, natural water evaporation is also a fundamental way to obtain fresh water for human beings. Here, a wafer-scale nanostructured silicon-based device that takes advantage of its well-aligned configuration that simultaneously realizes solar steam generation (SSG) for freshwater collection and hydrovoltaic effect generation for electricity output is developed. An ingenious porous, black carbon nanotube fabric (CNF) electrode endows the device with sustainable water self-pumping capability, excellent durable conductivity, and intense solar spectrum harvesting. A combined device based on the CNF electrode integrated with nanostructured silicon nanowire arrays (SiNWs) provided an aligned numerous surface-to-volume water evaporation interface that enables a recorded continuous short-circuit current 8.65 mA and a water evaporation rate of 1.31 kg m-2 h-1 under one sun illumination. Such wafer-scale SiNWs-based SSG and hydrovoltaic integration devices would unchain the bottleneck of the weak and discontinuous electrical output of hydrovoltaic devices, which inspires other sorts of semiconductor-based hydrovoltaic device designs to target superior performance.

Outcome and prognostic factors of CBF pediatric AML patients with t(8;21) differ from patients with inv(16).

PURPOSE: To explore the outcome and prognostic factors between inv(16) and t(8;21) disrupt core binding factor (CBF) in acute myeloid leukemia (AML). METHODS: The clinical characteristic, probability of achieving complete remission (CR), overall survival (OS) and cumulative incidence of relapse (CIR) were compared between inv(16) and (8;21). RESULTS: The CR rate was 95.2%, 10-year OS was 84.4% and CIR was 29.4%. Subgroup analysis showed that patients with t(8;21) had significant lower 10-year OS and CIR than patients with inv(16). Unexpectedly, there was a trend for pediatric AML receiving five courses cytarabine to have a lower CIR than four courses cytarabine (19.8% vs 29.3%, P = 0.06). Among the cohort of no-gemtuzumab ozogamicin(GO) treatment, inv (16) patients showed a similar 10-year OS (78.9% vs 83.5%; P = 0.69) and an inferior outcome on 10-year CIR (58.6% vs 28.9%, P = 0.01) than those patients with t(8;21). In contrast, inv (16) and t(8;21) patients receiving GO treatment had comparable OS (OS: 90.5% vs. 86.5%, P = 0.66) as well as CIR (40.4% vs. 21.4%, P = 0.13). CONCLUSION: Our data demonstrated that more cumulative cytarabine exposure could improve the outcome of childhood patients with t(8;21), while GO treatment was beneficial to the pediatric patients with inv(16).

Vincristine and dexamethasone pulses in addition to maintenance therapy among pediatric acute lymphoblastic leukemia (GD-ALL-2008): An open-label, multicentre, randomized, phase III clinical trial.

The efficacy and safety on the addition of vincristine (VCR) and dexamethasone (DEX) pulses to maintenance therapy among childhood acute lymphoblastic leukemia (ALL) remain uncertain. Herein, we perform an open-label, multicentre, randomized, phase III clinical trial that was conducted at nine major medical centers in Guangdong Province, China. Patients were randomly assigned either the conventional maintenance therapy (control group, n = 384) or the VCR/DEX pulse (treatment group, n = 375). When limited to the SR cohort, 10-year EFS was 82.6% (95% CI: 75.9-89.9) in the control group and 80.7% (95% CI: 74-88.1) in the treatment group (pnon-inferiority  = .0002). Similarly, patients with IR also demonstrated non-inferiority of the treatment group to the control group in terms of 10-year EFS (73.6% [95% CI: 67.6-80] vs. 77.6% [95% CI: 71.8-83.9]; pnon-inferiority  = .005). Among the HR cohort, compared with the control group, patients in the treatment group experienced a significant benefit in terms of 10-year EFS (61.1% [95% CI: 47.7-78.2] vs. 72.6% [95% CI: 55.6-94.7], p = .026) and a trend toward higher 10-year OS (73.8% [95% CI: 61.6-88.4] vs. 87.9% [95% CI: 579.2-97.5], p = .068). In the HR cohort, the total rate of drug-induced liver injury and Grade 3 chemotherapy-induced anemia were both lower for patients in the treatment group than in the control group (55.6% vs. 100%, p = .033; 37.5% vs. 60%, p = .036). Conversely, the total prevalence of chemotherapy-induced thrombocytopenia was higher for patients in the treatment group than in the control group (88.9% vs. 40%, p = .027). Pediatric acute lymphoblastic leukemia with high risk is suitable to VCR/DEX pulse during maintenance phase for the excellent outcome, while the standard-to-intermediate-risk patients could eliminate the pulses.

Inhibition of phosphorylated calcium/calmodulin-dependent protein kinase IIα relieves streptozotocin-induced diabetic neuropathic pain through regulation of P2X3 receptor in dorsal root ganglia.

Diabetic neuropathic pain (DNP) is frequent among patients with diabetes. We previously showed that P2X3 upregulation in dorsal root ganglia (DRG) plays a role in streptozotocin (STZ)-induced DNP but the underlying mechanism is unclear. Here, a rat model of DNP was established by a single injection of STZ (65 mg/kg). Fasting blood glucose was significantly elevated from the 1st to 3rd week. Paw withdrawal thresholds (PWTs) and paw withdrawal latencies (PWLs) in diabetic rats significantly reduced from the 2nd to 3rd week. Western blot analysis revealed that elevated p-CaMKIIα levels in the DRG of DNP rats were accompanied by pain-associated behaviors while CaMKIIα levels were unchanged. Immunofluorescence revealed significant increase in the proportion of p-CaMKIIα immune positive DRG neurons (stained with NeuN) in the 2nd and 3rd week and p-CaMKIIα was co-expressed with P2X3 in DNP rats. KN93, a CaMKII antagonist, significantly reduce mechanical hyperalgesia and thermal hyperalgesia and these effects varied dose-dependently, and suppressed p-CaMKIIα and P2X3 upregulation in the DRGs of DNP rats. These results revealed that the p-CaMKIIα upregulation in DRG is involved in DNP, which possibly mediated P2X3 upregulation, indicating CaMKIIα may be an effective pharmacological target for DNP management.

Dorsal root ganglia P2X4 and P2X7 receptors contribute to diabetes-induced hyperalgesia and the downregulation of electroacupuncture on P2X4 and P2X7.

Diabetic neuropathic pain (DNP) is highly common in diabetes patients. P2X receptors play critical roles in pain sensitization. We previously showed that elevated P2X3 expression in dorsal root ganglion (DRG) contributes to DNP. However, the role of other P2X receptors in DNP is unclear. Here, we established the DNP model using a single high-dose streptozotocin (STZ) injection and investigated the expression of P2X genes in the DRG. Our data revealed elevated P2X2, P2X4, and P2X7 mRNA levels in DRG of DNP rats. The protein levels of P2X4 and P2X7 in DNP rats increased, but the P2X2 did not change significantly. To study the role of P2X4 and P2X7 in diabetes-induced hyperalgesia, we treated the DNP rats with TNP-ATP (2',3'-O-(2,4,6-trinitrophenyl)-adenosine 5'-triphosphate), a nonspecific P2X1-7 antagonist, and found that TNP-ATP alleviated thermal hyperalgesia in DNP rats. 2 Hz electroacupuncture is analgesic against DNP and could downregulate P2X4 and P2X7 expression in DRG. Our findings indicate that P2X4 and P2X7 in L4-L6 DRGs contribute to diabetes-induced hyperalgesia, and that EA reduces thermal hyperalgesia and the expression of P2X4 and P2X7.

Analgesic effect of electroacupuncture on bone cancer pain in rat model: the role of peripheral P2X3 receptor.

Upregulation of P2X3 receptor (P2X3R) has been strongly implicated in nociceptive signaling including bone cancer pain (BCP). The present study, using rat bone cancer model, aimed to explore the role of P2X3R in regulating rat pain behavior under the intervention of electroacupuncture (EA). The BCP model was successfully established by injection with MRMT-1 breast cancer cell into the medullary cavity of left tibia for 3 × 104 cells/3 µL PBS in rats as revealed by obvious bone destruction, decreased paw withdrawal thresholds (PWTs), and reduced paw withdrawal latencies (PWLs). Western blot analyses showed that P2X3R expression was significantly upregulated in ipsilateral lumbar 4-6 (L4-6) dorsal root ganglia (DRG), but the difference not seen in spinal cord dorsal horn (SCDH). With the in-depth study of P2X3R activation, we observed that intrathecal injection of P2X3R agonist α,ß-meATP aggravated MRMT-1 induced BCP, while injection of P2X3R inhibitor A-317491 alleviated pain. Subsequently, we demonstrated that BCP induced mechanical allodynia and thermal hyperalgesia were attenuated after EA treatment. Under EA treatment, total P2X3R protein expression in ipsilateral DRGs was decreased, and it is worth mentioning that decreased expression of P2X3R membrane protein, which indicated that both the expression and membrane trafficking of P2X3R were inhibited by EA. The immunofluorescence assay showed that EA stimulation exerted functions by reducing the expression of P2X3R-positive cells in ipsilateral DRGs of BCP rats. Ca2+ imaging analysis revealed that the EA stimulation decreased the percentage of α,ß-meATP responsive neurons in DRGs and inhibited calcium influx. Notably, the inhibitory effect of EA on mechanical allodynia and nociceptive flinches was abolished by intrathecal injection of α,ß-meATP. These findings demonstrated EA stimulation ameliorated mechanical allodynia and thermal hyperalgesia in rat model of MRMT-1-induced BCP. EA exerts analgesic effect on BCP by reducing the overexpression and functional activity of P2X3R in ipsilateral DRGs of BCP rats. Our work first demonstrates the critical and overall role of P2X3R in EA's analgesia against peripheral sensitization of MRMT-1-induced BCP and further supports EA as a potential therapeutic option for cancer pain in clinic.

Electroacupuncture may alleviate diabetic neuropathic pain by inhibiting the microglia P2X4R and neuroinflammation.

Diabetic neuropathic pain (DNP) is a common and destructive complication of diabetes mellitus. The discovery of effective therapeutic methods for DNP is vitally imperative because of the lack of effective treatments. Although 2 Hz electroacupuncture (EA) was a successful approach for relieving DNP, the mechanism underlying the effect of EA on DNP is still poorly understood. Here, we established a rat model of DNP that was induced by streptozotocin (STZ) injection. P2X4R was upregulated in the spinal cord after STZ-injection. The upregulation of P2X4R was mainly expressed on activated microglia. Intrathecal injection of a P2X4R antagonist or microglia inhibitor attenuated STZ-induced nociceptive thermal hyperalgesia and reduced the overexpression of brain-derived neurotrophic factor (BDNF), interleukin-1ß (IL-1ß) and tumor necrosis factor-α (TNF-α) in the spinal cord. We also assessed the effects of EA treatment on the pain hypersensitivities of DNP rats, and further investigated the possible mechanism underlying the analgesic effect of EA. EA relieved the hyperalgesia of DNP. In terms of mechanism, EA reduced the upregulation of P2X4R on activated microglia and decreased BDNF, IL-1ß and TNF-α in the spinal cord. Mechanistic research of EA's analgesic impact would be beneficial in ensuring its prospective therapeutic effect on DNP as well as in extending EA's applicability.

Regulation of toll-like receptor (TLR) signaling pathways in atherosclerosis: from mechanisms to targeted therapeutics.

Atherosclerosis, one of the life-threatening cardiovascular diseases (CVDs), has been demonstrated to be a chronic inflammatory disease, and inflammatory and immune processes are involved in the origin and development of the disease. Toll-like receptors (TLRs), a class of pattern recognition receptors that trigger innate immune responses by identifying pathogen-associated molecular patterns (PAMPs) and danger-associated molecular patterns (DAMPs), regulate numerous acute and chronic inflammatory diseases. Recent studies reveal that TLRs have a vital role in the occurrence and development of atherosclerosis, including the initiation of endothelial dysfunction, interaction of various immune cells, and activation of a number of other inflammatory pathways. We herein summarize some other inflammatory signaling pathways, protein molecules, and cellular responses associated with TLRs, such as NLRP3, Nrf2, PCSK9, autophagy, pyroptosis and necroptosis, which are also involved in the development of AS. Targeting TLRs and their regulated inflammatory events could be a promising new strategy for the treatment of atherosclerotic CVDs. Novel drugs that exert therapeutic effects on AS through TLRs and their related pathways are increasingly being developed. In this article, we comprehensively review the current knowledge of TLR signaling pathways in atherosclerosis and actively seek potential therapeutic strategies using TLRs as a breakthrough point in the prevention and therapy of atherosclerosis.