Enzymes in Synergy: Bacteria Specific Molecular Probe for Locoregional Imaging of Urinary Tract Infection in vivo.

Uropathogenic Escherichia coli (UPECs) is a leading cause for urinary tract infections (UTI), accounting for 70-90% of community or hospital-acquired bacterial infections owing to high recurrence, imprecision in diagnosis and management, and increasing prevalence of antibiotic resistance. Current methods for clinical UPECs detection still rely on labor-intensive urine cultures that impede rapid and accurate diagnosis for timely UTI therapeutic management. Herein, we developed a first-in-class near-infrared (NIR) UPECs fluorescent probe (NO-AH) capable of specifically targeting UPECs through its collaborative response to bacterial enzymes, enabling locoregional imaging of UTIs both in vitro and in vivo. Our NO-AH probe incorporates a dual protease activatable moiety, which first reacts with OmpT, an endopeptidase abundantly present on outer membrane of UPECs, releasing an intermediate amino acid residue conjugated with a NIR hemicyanine fluorophore. Such liberated fragment would be subsequently recognized by aminopeptidase (APN) within periplasm of UPECs, activating localized fluorescence for precise imaging of UTIs in complex living environments. The peculiar specificity and selectivity of NO-AH, facilitated by the collaborative action of bacterial enzymes, features a timely and accurate identification of UPECs-infected UTIs, which could overcome misdiagnosis in conventional urine tests, thus opening new avenues towards reliable UTI diagnosis and personalized antimicrobial therapy management.

Acupuncture therapy on myofascial pain syndrome: a systematic review and meta-analysis.

Purpose: Traditional Chinese medicine (TCM) therapies, especially acupuncture, have received increasing attention in the field of pain management. This meta-analysis evaluated the effectiveness of acupuncture in the treatment of myofascial pain syndrome. Methods: A comprehensive search was conducted across a number of databases, including PubMed, Cochrane Library, WOS, CNKI, WANFANG, Sinomed, and VIP. Furthermore, articles of studies published from the inception of these databases until November 22, 2023, were examined. This systematic review and meta-analysis encompassed all randomized controlled trials (RCTs) on acupuncture for myofascial pain syndromes, without language or date restrictions. Based on the mean difference (MD) of symptom change, we critically assessed the outcomes reported in these trials. The quality of evidence was assessed using the Cochrane Risk of Bias Tool. The study is registered with PROSPERO under registration number CRD42023484933. Results: Our analysis included 10 RCTs in which 852 patients were divided into two groups: an acupuncture group (427) and a control group (425). The results of the study showed that acupuncture was significantly more effective than the control group in treating myofascial pain syndromes, which was reflected in a greater decrease in VAS scores (MD = -1.29, 95% [-1.65, -0.94], p < 0.00001). In addition, the improvement in PRI and PPI was more pronounced in the acupuncture group (PRI: MD = -2.04, 95% [-3.76, -0.32], p = 0.02) (PPI: MD = -1.03, 95% [-1.26, -0.79], p < 0.00001) compared to the control group. These results suggest that acupuncture is effective in reducing myofascial pain. It is necessary to further study the optimal acupoints and treatment time to achieve the best therapeutic effect. Systematic review registration: https://www.crd.york.ac.uk/prospero/, identifier CRD42023484933.

Boosting humoral and cellular immunity with enhanced STING activation by hierarchical mesoporous metal-organic framework adjuvants.

Vaccination is essential for preventing and controlling infectious diseases, along with reducing mortality. Developing safe and versatile adjuvants to enhance humoral and cellular immune responses to vaccines remains a key challenge in vaccine development. Here, we designed hierarchical mesoporous MOF-801 (HM801) using a Cocamidopropyl betaine (CAPB) and a Pluronics F127 in an aqueous phase system. Meanwhile, we synthesized a novel SARS-CoV-2 nanovaccine (R@M@HM801) with a high loading capacity for both the STING agonist (MSA-2) and the Delta receptor binding domain (Delta-RBD) antigen. R@M@HM801 enhanced MSA-2 and RBD utilization and effectively co-delivered MSA-2 and RBD antigens to antigen-presenting cells in the draining lymph nodes, thereby promoting the activation of both T and B cells. Lymphocyte single-cell analysis showed that R@M@HM801 stimulated robust CD11b+CD4+ T cells, CXCR5+CD4+ T follicular helper (Tfh), and durable CD4+CD44+CD62L-, CD8+CD44+CD62L- effector memory T cell (TEM) immune responses, and promoted the proliferative activation of CD26+ B cells in vivo. Meanwhile, R@M@HM801 induced stronger specific antibodies and neutralization of pseudovirus against Delta compared to the RBD + MAS-2 and RBD + MAS-2 + Alum vaccines. Our study demonstrated the efficacy of a hierarchical mesoporous HM801 and its potential immune activation mechanism in enhancing adaptive immune responses against viruses and other diseases.

Effects of dietary phytosterols or phytosterol esters supplementation on growth performance, biochemical blood indices and intestinal flora of C57BL/6 mice.

This study was conducted to evaluate the effects of phytosterols (PS) and phytosterol esters (PSE) on C57BL/6 mice. Three groups of 34 six-week-old C57BL/6 mice of specific pathogen free (SPF) grade, with an average initial body weight (IBW) of 17.7g, were fed for 24 days either natural-ingredient diets without supplements or diets supplemented with 89 mg/kg PS or diets supplemented with 400 mg/kg PSE. Growth performance, blood biochemistry, liver and colon morphology as well as intestinal flora status were evaluated. Both PS and PSE exhibited growth promotion and feed digestibility in mice. In blood biochemistry, the addition of both PS and PSE to the diet resulted in a significant decrease in Total Cholesterol (TC) and Triglyceride (TG) levels and an increase in Superoxide Dismutase (SOD) activity. No significant changes in liver and intestinal morphology were observed. Both increased the level of Akkermansia in the intestinal tract of mice. There was no significant difference between the effects of PS and PSE. It was concluded that dietary PS and PSE supplementation could improve growth performance, immune performance and gut microbiome structure in mice, providing insights into its application as a potential feed additive in animals production.

Shape Matters: Impact of Mesoporous Silica Nanoparticle Morphology on Anti-Tumor Efficacy.

A nanoparticle's shape is a critical determinant of its biological interactions and therapeutic effectiveness. This study investigates the influence of shape on the performance of mesoporous silica nanoparticles (MSNs) in anticancer therapy. MSNs with spherical, rod-like, and hexagonal-plate-like shapes were synthesized, with particle sizes of around 240 nm, and their other surface properties were characterized. The drug loading capacities of the three shapes were controlled to be 47.46%, 49.41%, and 46.65%, respectively. The effects of shape on the release behaviors, cellular uptake mechanisms, and pharmacological behaviors of MSNs were systematically investigated. Through a series of in vitro studies using 4T1 cells and in vivo evaluations in 4T1 tumor-bearing mice, the release kinetics, cellular behaviors, pharmacological effects, circulation profiles, and therapeutic efficacy of MSNs were comprehensively assessed. Notably, hexagonal-plate-shaped MSNs loaded with PTX exhibited a prolonged circulation time (t1/2 = 13.59 ± 0.96 h), which was approximately 1.3 times that of spherical MSNs (t1/2 = 10.16 ± 0.38 h) and 1.5 times that of rod-shaped MSNs (t1/2 = 8.76 ± 1.37 h). This research underscores the significance of nanoparticles' shapes in dictating their biological interactions and therapeutic outcomes, providing valuable insights for the rational design of targeted drug delivery systems in cancer therapy.

Venetoclax acts as an immunometabolic modulator to potentiate adoptive NK cell immunotherapy against leukemia.

Natural killer (NK) cell-based immunotherapy holds promise for cancer treatment; however, its efficacy remains limited, necessitating the development of alternative strategies. Here, we report that venetoclax, an FDA-approved BCL-2 inhibitor, directly activates NK cells, enhancing their cytotoxicity against acute myeloid leukemia (AML) both in vitro and in vivo, likely independent of BCL-2 inhibition. Through comprehensive approaches, including bulk and single-cell RNA sequencing, avidity measurement, and functional assays, we demonstrate that venetoclax increases the avidity of NK cells to AML cells and promotes lytic granule polarization during immunological synapse (IS) formation. Notably, we identify a distinct CD161lowCD218b+ NK cell subpopulation that exhibits remarkable sensitivity to venetoclax treatment. Furthermore, venetoclax promotes mitochondrial respiration and ATP synthesis via the NF-κB pathway, thereby facilitating IS formation in NK cells. Collectively, our findings establish venetoclax as a multifaceted immunometabolic modulator of NK cell function and provide a promising strategy for augmenting NK cell-based cancer immunotherapy.

Research on Medical Image Segmentation Method Based on Improved U-Net3.

Computer assisted diagnostic technology has been widely used in clinical practice, specifically focusing on medical image segmentation. Its purpose is to segment targets with certain special meanings in medical images and extract relevant features, providing reliable basis for subsequent clinical diagnosis and research. However, because of different shapes and complex structures of segmentation targets in different medical images, some imaging techniques have similar characteristics, such as intensity, color, or texture, for imaging different organs and tissues. The localization and segmentation of targets in medical images remains an urgent technical challenge to be solved. As such, an improved full scale skip connection network structure for the CT liver image segmentation task is proposed. This structure includes a biomimetic attention module between the shallow encoder and the deep decoder, and the feature fusion proportion coefficient between the two is learned to enhance the attention of the overall network to the segmented target area. In addition, based on the traditional point sampling mechanism, an improved point sampling strategy is proposed for characterizing medical images to further enhance the edge segmentation effect of CT liver targets. The experimental results on the commonly used combined (CT-MR) health absolute organ segmentation (CHAOS) dataset show that the average dice similarity coefficient (DSC) can reach 0.9467, the average intersection over union (IOU) can reach 0.9623, and the average F1 score can reach 0.9351. This indicates that the model can effectively learn image detail features and global structural features, leading to improved segmentation of liver images.

Deciphering the dual roles of PHD finger proteins from oncogenic drivers to tumor suppressors.

PHD (plant homeodomain) finger proteins emerge as central epigenetic readers and modulators in cancer biology, orchestrating a broad spectrum of cellular processes pivotal to oncogenesis and tumor suppression. This review delineates the dualistic roles of PHD fingers in cancer, highlighting their involvement in chromatin remodeling, gene expression regulation, and interactions with cellular signaling networks. PHD fingers' ability to interpret specific histone modifications underscores their influence on gene expression patterns, impacting crucial cancer-related processes such as cell proliferation, DNA repair, and apoptosis. The review delves into the oncogenic potential of certain PHD finger proteins, exemplified by PHF1 and PHF8, which promote tumor progression through epigenetic dysregulation and modulation of signaling pathways like Wnt and TGFß. Conversely, it discusses the tumor-suppressive functions of PHD finger proteins, such as PHF2 and members of the ING family, which uphold genomic stability and inhibit tumor growth through their interactions with chromatin and transcriptional regulators. Additionally, the review explores the therapeutic potential of targeting PHD finger proteins in cancer treatment, considering their pivotal roles in regulating cancer stem cells and influencing the immune response to cancer therapy. Through a comprehensive synthesis of current insights, this review underscores the complex but promising landscape of PHD finger proteins in cancer biology, advocating for further research to unlock novel therapeutic avenues that leverage their unique cellular roles.

Characterizing an effective magnetic field during asymmetric creep motion of Dzyaloshinskii domain walls.

Understanding the role of Landau Lifshitz Gilbert dynamics in describing magnetic domain wall (DW) motion in the creep regime is complicated by the presence of static pinning, but has regained interest due to recently observed directional growth in thin films with significant interfacial Dzyaloshinskii-Moriya interaction. Here, we delve into this directional domain growth behaviour in Pt/Co/Ni-based multi-layers under the influence of combined longitudinal and perpendicular magnetic fields via magneto-optical Kerr effect microscopy. Observations, including the onset field,µ0Honset, where the growth direction reverses by 180 degrees, align with the transient steady-state model predictions. By systematically varying the applied perpendicular magnetic field, we estimate the strength of an effective perpendicular field that acts on the DW during creep movement, which was expectedly found to be much smaller than the applied external field itself. This work further adds to the complexity of asymmetric domain expansion in the creep regime, but also highlights the range of magnetic information that can be extracted from careful analysis of this behavior.

Therapeutic efficacy of acupuncture point stimulation for stomach cancer pain: a systematic review and meta-analysis.

Purpose: In recent years, traditional Chinese medicine has received widespread attention in the field of cancer pain treatment. This meta-analysis is the first to evaluate the effectiveness and safety of acupuncture point stimulation in the treatment of stomach cancer pain. Methods: For this systematic review and meta-analysis, we searched PubMed, Web of Science, Cochrane Library, Embase, WANFANG, China National Knowledge Infrastructure (CNKI), and Chinese Journal of Science and Technology (VIP) databases as well as forward and backward citations to studies published between database creation to July 27, 2023. All randomized controlled trials (RCTs) on acupuncture point stimulation for the treatment of patients with stomach cancer pain were included without language restrictions. We assessed all outcome indicators of the included trials. The evidence from the randomized controlled trials was synthesized as the standardized mean difference (SMD) of symptom change. The quality of the evidence was assessed using the Cochrane Risk of Bias tool. This study is registered on PROSPERO under the number CRD42023457341. Results: Eleven RCTs were included. The study included 768 patients, split into 2 groups: acupuncture point stimulation treatment group (n = 406), medication control group (n = 372). The results showed that treatment was more effective in the acupuncture point stimulation treatment group than in the medication control group (efficacy rate, RR = 1.63, 95% CI 1.37 to 1.94, p < 0.00001), decreasing in NRS score was greater in acupuncture point stimulation treatment group than in the medication control group (SMD = -1.30, 95% CI -1.96 to -0.63, p < 0.001). Systematic Review Registration: https://clinicaltrials.gov/, identifier CRD42023457341.

Hierarchical joint analysis of marginal summary statistics-Part I: Multipopulation fine mapping and credible set construction.

Recent advancement in genome-wide association studies (GWAS) comes from not only increasingly larger sample sizes but also the shift in focus towards underrepresented populations. Multipopulation GWAS increase power to detect novel risk variants and improve fine-mapping resolution by leveraging evidence and differences in linkage disequilibrium (LD) from diverse populations. Here, we expand upon our previous approach for single-population fine-mapping through Joint Analysis of Marginal SNP Effects (JAM) to a multipopulation analysis (mJAM). Under the assumption that true causal variants are common across studies, we implement a hierarchical model framework that conditions on multiple SNPs while explicitly incorporating the different LD structures across populations. The mJAM framework can be used to first select index variants using the mJAM likelihood with different feature selection approaches. In addition, we present a novel approach leveraging the ideas of mediation to construct credible sets for these index variants. Construction of such credible sets can be performed given any existing index variants. We illustrate the implementation of the mJAM likelihood through two implementations: mJAM-SuSiE (a Bayesian approach) and mJAM-Forward selection. Through simulation studies based on realistic effect sizes and levels of LD, we demonstrated that mJAM performs well for constructing concise credible sets that include the underlying causal variants. In real data examples taken from the most recent multipopulation prostate cancer GWAS, we showed several practical advantages of mJAM over other existing multipopulation methods.

Predicting delayed extubation and transfer to the intensive care unit in children undergoing posterior fusion surgery for scoliosis : A retrospective observational study.

BACKGROUND: Delayed extubation and transfer to the intensive care unit (ICU) in children undergoing major scoliosis surgery may increase postoperative complications, prolong hospital stay, and increase medical expenses; however, whether a child will require delayed extubation or transfer to the ICU after scoliosis orthopedic surgery is not fully understood. In this study, we reviewed the risk factors for delayed extubation and transfer to the ICU after scoliosis orthopedic surgery in children. METHOD: The electronic medical records of pediatric patients (≤ 18 years) who underwent posterior spinal fusion surgery between January 2018 and November 2021 were reviewed and analyzed. Patient characteristics (age, sex, body mass index, American Society of Anesthesiologists, ASA, grade, preoperative lung function, and congenital heart disease), preoperative Cobb angle, scoliosis type, correction rate, vertebral fusion segments, pedicle screws, surgical osteotomy, intraoperative bleeding, intraoperative allogeneic transfusion, intraoperative hemoglobin changes, intraoperative mean arterial pressure changes, intraoperative tidal volume (ml/kg predicted body weight), surgical time, postoperative extubation, and transfer to the ICU were collected. The primary outcomes were delayed extubation and transfer to the ICU. Multivariate logistic regression models were used to determine the risk factors for delayed extubation and ICU transfer. RESULTS: A total of 246 children who satisfied the inclusion criteria were enrolled in this study, of whom 23 (9.3%) had delayed extubation and 81 (32.9%) were transferred to the ICU after surgery. High ASA grade (odds ratio [OR] 5.42; 95% confidence interval [CI] 1.49-19.78; p = 0.010), high Cobb angle (OR 1.04; 95% CI 1.02-1.07; p < 0.001), moderate to severe pulmonary dysfunction (OR 10.9; 95% CI 2.00-59.08; p = 0.006) and prolonged surgical time (OR 1.01; 95% CI 1.00-1.03; p = 0.040) were risk factors for delayed extubation. A high Cobb angle (OR 1.02; 95% CI 1.01-1.04; p = 0.004), high intraoperative bleeding volume (OR 1.06; 95% CI 1.03-1.10; p = 0.001), allogeneic transfusion (OR 3.30; 95% CI 1.24-8.83; p = 0.017) and neuromuscular scoliosis (OR 5.38; 95% CI 1.59-18.25; p = 0.007) were risk factors for transfer to the ICU. A high Cobb angle was a risk factor for both delayed extubation and ICU transfer. Age, sex, body mass index, number of vertebral fusion segments, correction rate, and intraoperative tidal volume were not associated with delayed postoperative extubation and ICU transfer. CONCLUSION: The most common risk factor for delayed extubation and ICU transfer in pediatric patients who underwent posterior spinal fusion was a high Cobb angle. Determining risk factors for a poor prognosis may help optimize perioperative respiratory management strategies and planning of postoperative care for children undergoing complicated spinal surgery.

Codetection of Proteins and RNAs on Extracellular Vesicles for Pancreatic Cancer Early Diagnosis.

Tumor-derived extracellular vesicles (EVs) carry tumor-specific proteins and RNAs, thus becoming prevalent targets for early cancer diagnosis. However, low expression of EV cargos and insufficient diagnostic power of individual biomarkers hindered EVs application in clinical practice. Herein, we propose a multiplex Codetection platform of proteins and RNAs (Co-PAR) for EVs. Co-PAR adopted a pair of antibody-DNA probes to recognize the same target protein, which in turn formed a double-stranded DNA. Thus, the target protein could be quantified by detecting the double-stranded DNA via qPCR. Meanwhile, qRT-PCR simultaneously quantified the target RNAs. Thus, with a regular qPCR instrument, Co-PAR enabled the codetection of multiplex proteins and RNAs, with the sensitivity of 102 EVs/µL (targeting CD63) and 1 EV/µL (targeting snRNA U6). We analyzed the coexpressions of three protein markers (CD63, GPC-1, HER2) and three RNA markers (snRNA U6, GPC-1 mRNA, miR-10b) on EVs from three pancreatic cell lines and 30 human plasma samples using Co-PAR. The diagnostic accuracy of the 6-biomarker combination reached 92.9%, which was at least 6.2% higher than that of 3-biomarker combinations and at least 13.5% higher than that of 6 single biomarkers. Co-PAR, as a multiparameter detection platform for EVs, has great potential in early disease diagnosis.

Robust Tensor-Based DOA and Polarization Estimation in Conformal Polarization Sensitive Array with Bad Data.

Partially impaired sensor arrays pose a significant challenge in accurately estimating signal parameters. The occurrence of bad data is highly probable, resulting in random loss of source information and substantial performance degradation in parameter estimation. In this paper, a tensor variational sparse Bayesian learning (TVSBL) method is proposed for the estimate of direction of arrival (DOA) and polarization parameters jointly based on a conformal polarization sensitive array (CPSA), taking into account scenarios with the partially impaired sensor array. First, a sparse tensor-based received data model is developed for CPSAs that incorporates bad data. Then, a column vector detection method is proposed to diagnose the positions of the impaired sensors. In scenarios involving partially impaired sensor arrays, a low-rank matrix completion method is employed to recover the random loss of signal information. Finally, variational sparse Bayesian learning (VSBL) and minimum eigenvector methods are utilized sequentially to obtain the DOA and polarization parameters estimation, successively. Furthermore, the Cramér-Rao bound is given for the proposed method. Simulation results validated the effectiveness of the proposed method.

Endothelial RSPO3 mediates pulmonary endothelial regeneration by LGR4-dependent activation of ß-catenin and ILK signaling pathways after inflammatory vascular injury.

Endothelial repair is essential for restoring tissue fluid homeostasis following lung injury. R-spondin3 (RSPO3), a secreted protein mainly produced by endothelial cells (ECs), has shown its protective effect on endothelium. However, the specific mechanisms remain unknown. To explore whether and how RSPO3 regulates endothelial regeneration after inflammatory vascular injury, the role of RSPO3 in sepsis-induced pulmonary endothelial injury was investigated in EC-specific RSPO3 knockdown, inducible EC-specific RSPO3 deletion mice, EC-specific RSPO3 overexpression mice, systemic RSPO3-administration mice, in isolated mouse lung vascular endothelial cells (MLVECs), and in plasma from septic patients. Here we show that plasma RSPO3 levels are decreased in septic patients and correlated with endothelial injury markers and PaO2/FiO2 index. Both pulmonary EC-specific knockdown of RSPO3 and inducible EC-specific RSPO3 deletion inhibit pulmonary ECs proliferation and exacerbate ECs injury, whereas intra-pulmonary EC-specific RSPO3 overexpression promotes endothelial recovery and attenuates ECs injury during endotoxemia. We show that RSPO3 mediates pulmonary endothelial regeneration by a LGR4-dependent manner. Except for ß-catenin, integrin-linked kinase (ILK)/Akt is also identified as a novel downstream effector of RSPO3/LGR4 signaling. These results conclude that EC-derived RSPO3 mediates pulmonary endothelial regeneration by LGR4-dependent activation of ß-catenin and ILK signaling pathways after inflammatory vascular injury.

Duration-specific association between plasma IGFBP7 levels and diabetic complications in patients with type 2 diabetes mellitus.

OBJECTIVE: Insulin-like growth factor binding protein 7 (IGFBP7) has a strong affinity to insulin. This study aimed to evaluate the relationship between IGFBP7 and complications among type 2 diabetes mellitus (T2DM) patients. DESIGN: A total of 1449 T2DM patients were selected from a cross-sectional study for disease management registered in the National Basic Public Health Service in Changshu, China, and further tested for their plasma IGFBP7 levels. Logistic regressions and Spearman's rank correlation analyses were used to explore the associations of IGFBP7 with diabetic complications and clinical characteristics, respectively. RESULTS: Among the 1449 included T2DM patients, 403 (27.81%) had complications. In patients with shorter duration (less than five years), the base 10 logarithms of IGFBP7 concentration were associated with T2DM complications, with an adjusted odds ratio (OR) of 2.41 [95% confidence interval (95%CI) = 1.06-5.48]; while in patients with longer duration (more than five years), plasma IGFBP7 levels were not associated with T2DM complications. Furthermore, in T2DM patients with shorter duration, those with two or more types of complications were more likely to have higher levels of IGFBP7. CONCLUSION: IGFBP7 is positively associated with the risk of complication in T2DM patients with shorter duration.

SpiDe-Sr: blind super-resolution network for precise cell segmentation and clustering in spatial proteomics imaging.

Spatial proteomics elucidates cellular biochemical changes with unprecedented topological level. Imaging mass cytometry (IMC) is a high-dimensional single-cell resolution platform for targeted spatial proteomics. However, the precision of subsequent clinical analysis is constrained by imaging noise and resolution. Here, we propose SpiDe-Sr, a super-resolution network embedded with a denoising module for IMC spatial resolution enhancement. SpiDe-Sr effectively resists noise and improves resolution by 4 times. We demonstrate SpiDe-Sr respectively with cells, mouse and human tissues, resulting 18.95%/27.27%/21.16% increase in peak signal-to-noise ratio and 15.95%/31.63%/15.52% increase in cell extraction accuracy. We further apply SpiDe-Sr to study the tumor microenvironment of a 20-patient clinical breast cancer cohort with 269,556 single cells, and discover the invasion of Gram-negative bacteria is positively correlated with carcinogenesis markers and negatively correlated with immunological markers. Additionally, SpiDe-Sr is also compatible with fluorescence microscopy imaging, suggesting SpiDe-Sr an alternative tool for microscopy image super-resolution.

Photosensitive Hydrogel with Temperature-Controlled Reversible Nano-Apertures for Single-Cell Protein Analysis.

Single cell western blot (scWB) is one of the most important methods for cellular heterogeneity profiling. However, current scWB based on conventional photoactive polyacrylamide hydrogel material suffers from the tradeoff between in-gel probing and separation resolution. Here, a highly sensitive temperature-controlled single-cell western blotting (tc-scWB) method is introduced, which is based on a thermo/photo-dualistic-sensitive polyacrylamide hydrogel, namely acrylic acid-functionalized graphene oxide (AFGO) assisted, N-isopropylacrylamide modified polyacrylamide (ANP) hydrogel. The ANP hydrogel is contracted at high-temperature to constrain protein band diffusion during microchip electrophoretic separation, while the gel aperture is expanded under low-temperature for better antibody penetration into the hydrogel. The tc-scWB method enables the separation and profiling of small-molecule-weight proteins with highly crosslinked gel (12% T) in SDS-PAGE. The tc-scWB is demonstrated on three metabolic and ER stress-specific proteins (CHOP, MDH2 and FH) in four pancreatic cell subtypes, revealing the expression of key enzymes in the Krebs cycle is upregulated with enhanced ER stress. It is found that ER stress can regulate crucial enzyme (MDH2 and FH) activities of metabolic cascade in cancer cells, boosting aerobic respiration to attenuate the Warburg effect and promote cell apoptosis. The tc-scWB is a general toolbox for the analysis of low-abundance small-molecular functional proteins at the single-cell level.