High Mobility Group Protein B1 Levels Research Articles

The Ability of Clinically Relevant Chemotherapeutics to Induce Immunogenic Cell Death in Squamous Cell Carcinoma.

Immunogenic cell death (ICD) is a crucial mechanism for triggering the adaptive immune response in cancer patients. Damage-associated molecular patterns (DAMPs) are critical factors in the detection of ICD. Chemotherapeutic drugs can cause ICD and the release of DAMPs. The aim of this study was to assess the potential for paclitaxel and platinum-based chemotherapy regimens to induce ICD in squamous cell carcinoma (SCC) cell lines. In addition, we examined the immunostimulatory effects of clinically relevant chemotherapeutic regimens utilized in the treatment of SCC. We screened for differentially expressed ICD markers in the supernatants of three SCC cell lines following treatment with various chemotherapeutic agents. The ICD markers included Adenosine Triphosphate (ATP), Calreticulin (CRT), Annexin A1 (ANXA 1), High Mobility Group Protein B1 (HMGB1), and Heat Shock Protein 70 (HSP70). A vaccination assay was also employed in C57BL/6J mice to validate our in vitro findings. Lastly, the levels of CRT and HMGB1 were evaluated in Serum samples from SCC patients. Addition of the chemotherapy drugs cisplatin (DDP), carboplatin (CBP), nedaplatin (NDP), oxaliplatin (OXA) and docetaxel (DOC) increased the release of ICD markers in two of the SCC cell lines. Furthermore, mice that received vaccinations with cervical cancer cells treated with DDP, CBP, NDP, OXA, or DOC remained tumor-free. Although CBP induced the release of ICD-associated molecules in vitro, it did not prevent tumor growth at the vaccination site in 40% of mice. In addition, both in vitro and in vivo results showed that paclitaxel (TAX) and LBP did not induce ICD in SCC cells. The present findings suggest that chemotherapeutic agents can induce an adjuvant effect leading to the extracellular release of DAMPs. Of the agents tested here, DDP, CBP, NDP, OXA and DOC had the ability to act as inducers of ICD.

HMGB1 in exosomes derived from gastric cancer cells induces M2-like macrophage polarization by inhibiting the NF-κB signaling pathway.

Gastric cancer (GC) seriously threatens human health. High mobility group protein B1 (HMGB1) and M2-like macrophages are closely associated with core events about human cancers, such as invasion, and metastasis, and cancer microenvironment. This study mainly determined the regulatory effect of HMGB1 in GC cell-derived exosomes on M2-like macrophage polarization as well as the underlying mechanism. HMGB1 was found to be highly expressed in gastric tissue specimens, which might lead to the poor prognosis of GC. High levels of HMGB1 were also observed in the plasma of GC patients, indicating the possibility that it regulates the immune microenvironment via exosomes. Further study revealed and confirmed the regulatory effect of exosomes derived from GC cells with high HMGB1 level on inducing M2-like macrophage polarization. Mechanistically, by interacting with the transcription factor POU2F1, exosomal HMGB1 inhibited the transcriptional activity of p50, resulting in the inactivation of NF-κB signaling pathway and thereby inducing M2-like macrophage polarization. Moreover, instead of promoting the proliferation of GC cells, exosomes with high HMGB1 levels induced M2-like macrophage polarization and promoted GC progression. This study reveals a novel mechanism by which HMGB1 promotes GC progression, which may provide new insights for improving the efficacy of cancer immunotherapy.

Honokiol Prevents Intestinal Barrier Dysfunction in Mice with Severe Acute Pancreatitis and Inhibits JAK/STAT1 Pathway and Acetylation of HMGB1.

To investigate the effect of honokiol (HON) and the role of high-mobility group protein B1 (HMGB1) on the pathogenesis of severe acute pancreatitis (SAP). Thirty mice were numbered according to weight, and randomly divided into 5 groups using a random number table, including control, SAP, SAP and normal saline (SAP+NS), SAP and ethyl pyruvate (SAP+EP), or SAP+HON groups, 6 mice in each group. Samples of pancreas, intestine, and blood were collected 12 h after SAP model induction for examination of pathologic changes, immune function alterations by enzyme linked immunosorbent assay (ELISA), and Western blot. In vitro experiments, macrophages were divided into 5 groups, the control, lipopolysaccharide (LPS), LPS+DMSO (DMSO), LPS+anti-HMGB1 monoclonal antibody (mAb), and LPS+ HON groups. The tight connection level was determined by transmission electron microscopy and fluorescein isothiocyanate-labeled. The location and acetylation of HMGB1 were measured by Western blot. Finally, pyridone 6 and silencing signal transducer and activator of the transcription 1 (siSTAT1) combined with honokiol were added to determine whether the Janus kinase (JAK)/ STAT1 participated in the regulation of honokiol on HMGB1. The protein expression levels of HMGB1, JAK, and STAT1 were detected using Western blot. Mice with SAP had inflammatory injury in the pancreas, bleeding of intestinal tissues, and cells with disrupted histology. Mice in the SAP+HON group had significantly fewer pathological changes. Mice with SAP also had significant increases in the serum levels of amylase, lipase, HMGB1, tumor necrosis factor- α, interleukin-6, diamine oxidase, endotoxin-1, and procalcitonin. Mice in the SAP+HON group did not show these abnormalities (P<0.01). Studies of Caco-2 cells indicated that LPS increased the levels of occludin and claudin-1 as well as tight junction permeability, decreased the levels of junctional adhesion molecule C, and elevated intercellular permeability (P<0.01). HON treatment blocked these effects. Studies of macrophages indicated that LPS led to low nuclear levels of HMGB1, however, HON treatment increased the nuclear level of HMGB1 (P<0.01). HON treatment also inhibited the expressions of JAK1, JAK2, and STAT1 (P<0.01) and increased the acetylation of HMGB1 (P<0.05). HON prevented intestinal barrier dysfunction in SAP by inhibiting HMGB1 acetylation and JAK/STAT1 pathway.

The correlation between the severity of cerebral microbleeds and serum HMGB1 levels and cognitive impairment in patients with cerebral small vessel disease.

The study investigated the correlation and predictive value between the severity of cerebral microbleeds (CMBs) and the level of serum High Mobility Group Protein B1 (HMGB1) and the occurrence of cognitive impairment in patients with cerebral small vessel disease (CSVD). A total of 139 patients with CSVD admitted to the Department of Neurology of the First Affiliated Hospital of Xinxiang Medical University from December 2020 to December 2022 were selected as study subjects. The Montreal Cognitive Assessment (MoCA) scale was used to assess the cognitive function and was divided into the cognitive impairment group and the cognitive normal group. Magnetic Resonance Imaging (MRI) and Susceptibility Weighted Imaging (SWI) were used to screen and assess the severity of CMBs. Serum HMGB1 levels of CSVD patients were measured by enzyme linked immunosorbent assay (ELISA). Multivariable logistic regression analysis was used to explore risk factors for cognitive impairment and CMBs. Pearson correlation analysis was used to investigate the correlation between HMGB1 and cognitive function. Receiver Operating Characteristics (ROC) curves were used to assess the predictive value of HMGB1 for the occurrence of cognitive impairment in patients with CMBs. High Mobility Group Protein B1, uric acid (UA), glycosylated hemoglobin (HbA1c), CMBs, lacunar cerebral infarction (LI), years of education, and history of hypertension were risk factors for cognitive impairment (P < 0.05); HMGB1 was significantly and negatively associated with total MoCA score, visuospatial/executive ability, and delayed recall ability (P < 0.05). HMGB1 was significantly and positively correlated with the number of CMBs (P < 0.05). The area under the ROC curve for HMGB1 predicting cognitive impairment in patients with CMBs was 0.807 (P < 0.001). Serum HMGB1 levels are associated with the development of cognitive impairment in CSVD patients, and serum HMGB1 levels have a high predictive value for the development of cognitive impairment in CSVD patients with combined CMBs, which can be used for early clinical identification and intervention of vascular cognitive impairment.

Regulation of RIP1-Mediated necroptosis via necrostatin-1 in periodontitis.

To explore the mechanism of receptor-interacting protein 1 (RIP1)-mediated necroptosis during periodontitis progression. RIP3 and mixed lineage kinase domain-like protein (MLKL) have been detected to be upregulated in periodontitis models. Because RIP1 is involved in necroptosis, it might also play a role in the progression of periodontitis. An experimental periodontitis model in BALB/c mice was established by inducing oral bacterial infection. Western blotting and immunofluorescence analyses were used to detect RIP1 expression in the periodontal ligament. Porphyromonas gingivalis was used to stimulate L929 and MC3T3-E1. RIP1 was inhibited using small-interfering RNA. Western blotting, reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and enzyme-linked immunosorbent assay (ELISA) analyses were used to detect the effect of necroptosis inhibition on the expression of damage-associated molecular patterns and inflammatory cytokines. Necrostatin-1 (Nec-1) was intraperitoneally injected to inhibit RIP1 expression in mice. Necroptosis activation and inflammatory cytokine expression in periodontal tissue were verified. Tartrate-resistant acid phosphatase staining was applied to observe osteoclasts in the bone tissues of different groups. RIP1-mediated necroptosis was activated in mice with periodontitis. P. gingivalis induced RIP1-mediated necroptosis in L929 and MC3T3-E1 cells. After RIP1 inhibition, the expression levels of high mobility group protein B1 (HMGB1) and inflammatory cytokines were downregulated. After inhibiting RIP1 with Nec-1 in vivo, necroptosis was also inhibited, the expression levels of HMGB1 and inflammatory cytokines were downregulated, and osteoclast counts in the periodontal tissue decreased. RIP1-mediated necroptosis plays a role in the pathological process of periodontitis in mice. Nec-1 inhibited necroptosis, alleviated inflammation in periodontal tissue, and reduced bone resorption in periodontitis.

HMGB1 promotes the development of castration‑resistant prostate cancer by regulating androgen receptor activation.

AR signalling pathway reactivation plays a key role in the development of castration-resistant prostate cancer (CRPC). High-mobility group protein B1 (HMGB1) is an important factor involved in the occurrence and development of a variety of tumours by regulating gene transcription. In the present study, the association between HMGB1 and prostate cancer (PCa) and the effects of HMGB1 on androgen receptor (AR) transcription and signalling pathway reactivation in PCa cells in vitro and in vivo were evaluated. A bioinformatics method was used to determine the mRNA expression level of HMGB1 in PCa specimens and its correlation with the mRNA expression of AR. Immunohistochemical staining was used to detect the expression of these proteins in clinical PCa samples. Reporter gene and ChIP assays were performed to determine the activity of AR and the effect of HMGB1 on the ability of AR to bind to the promoters of prostate specific antigen and transmembrane protease, serine 2. A bioluminescence resonance energy transfer assay was employed to observe the direct interaction between HMGB1 and AR protein. Additionally, a castrated nude mouse xenograft tumour model was established to verify the effect of HMGB1 on PCa. The results revealed that HMGB1 expression was significantly increased in PCa specimens, which may have a strong correlation with AR expression. Moreover, HMGB1 could reactivate the AR signalling pathway, directly interact with AR, and promote the development of CRPC in an androgen-independent manner. The results of the present study indicated that HMGB1 promoted the development of CRPC by interacting with AR, which inferred that decreasing the expression of HMGB1 may be a potential effective method for CRPC prevention and treatment.

The Value of Serum HMGB1 and sRAGE in the Diagnosis, Efficacy Monitoring and Prognosis of Newly Diagnosed Multiple Myeloma

To evaluate the value of high mobility group protein B1 (HMGB1) and soluble receptor for advanced glycation end products (sRAGE) in the diagnosis, efficacy monitoring and prognosis of newly diagnosed multiple myeloma (MM) patients. Fifty newly diagnosed MM patients before and after chemotherapy and 50 hematological outpatients from October 2018 to May 2020 were selected. Enzyme linked immunosorbent assay (ELISA) was used to detect the serum HMGB1 and sRAGE levels of the patients. ROC was used to further analyze the efficacy of serum HMGB1 and sRAGE levels on the diagnosis of MM. At the same time, the serum levels of HMGB1 and sRAGE before and after chemotherapy were compared, and their values in the evaluation of curative effect of MM patients were analyzed. According to the mean values of serum HMGB1 and sRAGE, all the patients were divided into different groups, the clinical characteristics and survival status of the patients were compared. Before treatment the serum HMGB1 level of the patients in MM group was higher than that in control group, while sRAGE level was lower (t=11.363,6.127, P<0.001). The AUC of serum HMGB1 and sRAGE in the MM patients was 0.955 and 0.811, respectively. After 3 courses of chemotherapy, HMGB1 level of the patients in CR group was lower than before chemotherapy, while in PD group was higher, as well as sRAGE level of the patients in PR group (P<0.05). There were significant differences in R-ISS stage, HGB, CRP, ESR, CD56, CD117, D13S319 deletion between HMGB1 high expression group and HMGB1 low expression group (χ2=3.920, 6.522, 6.65, 4.16, 3.945, 6.65, 4.16, P<0.05), while there were significant differences in ISS stage, CRP and CD56 between sRAGE low expression group (28 cases) and sRAGE high expression group (22 cases) (χ2=4.565, 4.711, 5.547, P<0.05). Kaplan-Meier survival analysis showed that the patients in HMGB1 low expression group had better survival condition, for PFS Tlow>Thigh (χ2=9.470, P<0.05), and for OS Tlow>Thigh (χ2=7.808, P<0.05); there was no difference in the survival of sRAGE high expression group and low expression group, for PFS Tlow<Thigh (χ2=1.661, P>0.05), and for OS Tlow<Thigh (χ2=2.048, P>0.05). Cox analysis showed that LDH and HMGB1 were the factors affecting the prognosis of the patients, and both of them affected PFS (HR=2.771, 95% CI: 1.002-7.662, P=0.049; HR=6.022, 95% CI: 1.689-21.470, P=0.006), while HMGB1 also affected OS (HR=4.275, 95% CI: 1.183-15.451, P=0.027). The serum HMGB1 and sRAGE have certain auxiliary value for the diagnosis and curative effect monitoring of newly diagnosed MM patients, and serum HMGB1 is expected to be an auxiliary detection index for the prognosis of MM.

Effect of paeoniflorin on distal survival of random flaps

BackgroundDistal ischemic necrosisis a common complication of orthopedic random skin flaps surgery. Paeoniflorin, a natural compound extracted from Paeonia lactiflora, can enhances angiogenesis and alleviates excessive inflammatory response. We investigated the changes of ischemic extra-long flaps with paeoniflorin and its possible mechanism. MethodsWe raised dorsal McFarlane flaps in 54 Sprague-Dawley rats. We designed three groups of rats: high-paeoniflorin group (HP, 50 mg/kg/d), low-paeoniflorin group (LP, 20 mg/kg/d), and control group. The flap survival rate was calculated, seven days after flap construction.Blood perfusion was detected by laser Doppler flow imaging, and angiogenesis wasdetected by Lead oxide/gelatin angiography.Oxidative stress levels of flaps were determined by detecting superoxide dismutase (SOD) and malondialdehyde (MDA). The histopathological status of flap was evaluated by hematoxylin and eosin (H&E) staining.Immunohistochemistry was used to determine the expression of high mobility group protein B1 (HMGB1), nuclear factor-kappa B (NF-κB), Toll-like receptor 4 (TLR4), tumor necrosis factor-α (TNF-α), interleukin (IL)-6, IL-1β, IL-18, vascular endothelial growth factor (VEGF), cysteine protease-1 (caspase-1) and NLPR3. ResultsThe flap survival rates and SOD activity in the experimental groups were significantly higher, while MDA activity was lower. Experimental groups showed significantly improved microcirculatory blood flow to the flap and increased angiogenesis. Immunohistochemistry revealed that paeoniflorin was associated with significantly increased VEGF expression, and decreased level of HMGB1, TLR4, TNF-α, NF-κB, IL-6, IL-1β, caspase-1, NLPR3, and IL-18. ConclusionsPaeoniflorin effectively enhanced the survival of rat random skin flaps by promoting vascular hyperplasia, inhibiting pyroptosis, and down-regulating inflammation.

Sonosensitized Aggregation-Induced Emission Dots with Capacities of Immunogenic Cell Death Induction and Multivalent Blocking of Programmed Cell Death-Ligand 1 for Amplified Antitumor Immunotherapy

Sonosensitized Aggregation-Induced Emission Dots with Capacities of Immunogenic Cell Death Induction and Multivalent Blocking of Programmed Cell Death-Ligand 1 for Amplified Antitumor Immunotherapy

Changes of Inflammasome in Children with Immune Thrombocytopenia before and after Treatment

To clarify the significance of inflammasome NLRP3 in children with immune thrombocytopenia (ITP) by detecting its changes before and after treatment. Twenty children with ITP diagnosed and treated in Xuzhou Children's Hospital were enrolled as observation group, and 10 healthy children as control group. The mRNA levels of NLRP3, ASC, and Caspase-1 were measured by real-time quantitative PCR (RT-qPCR), the serum levels of IL-18, IL-1β, and high mobility group protein B1 (HMGB1) were detected by ELISA, and the protein level of NLRP3 was detected by Western blot. In newly diagnosed ITP children, the serum levels of IL-18, IL-1β and HMGB1 significantly decreased after treatment (P<0.05). After treatment, NLRP3, ASC, and Caspase-1 mRNA levels in peripheral blood mononuclear cells were significantly lower than those before treatment (P<0.05). NLRP3 protein expression decreased significantly after treatment. Expression of NLRP3 inflammasome and downstream inflammatory factors are decrease after treatment in children with ITP, which may be used as effective prognostic markers.

Matrix and Sampling Effects on Quantification of Protein Biomarkers of Drug-Induced Liver Injury.

Macrophage colony stimulating factor 1 receptor (MCSF1R), osteopontin (OPN), high-mobility group protein B1 (HMGB1), glutamate dehydrogenase (GLDH), keratin 18 (K18), and caspase-cleaved keratin 18 (ccK18) are considered promising mechanistic biomarkers for the diagnosis of drug-induced liver injury. Here, we aim to elucidate the impact of the sample matrix and handling on the quantification of these emerging protein biomarkers. We investigated effects such as time from collection to centrifugation during serum (± gel) or EDTA plasma preparation on two assay platforms: immunoaffinity liquid chromatography mass spectrometric assays and sandwich immunoassays. Furthermore, we measured GLDH activity with an enzymatic activity assay. Matrix effects were observed particularly for HMGB1 and MCSF1R. HMGB1 levels were higher in serum than in plasma, whereas higher concentrations of MCSF1R were observed in plasma than in serum. A comparison of sample collection to centrifugation time ranging from 15 to 60 min demonstrated increasing levels of HMGB1 in serum, while MCSF1R, OPN, GLDH, and ccK18 concentrations remained stable. Additionally, there was a poor correlation in HMGB1 and ccK18 levels between serum and plasma. Considering the observed matrix effects, we recommend plasma as a matrix of choice and cross-study comparison studies to be limited to those using the same matrix.

MiR-142-3p reduces the viability of human cervical cancer cells by negatively regulating the cytoplasmic localization of HMGB1.

High mobility group protein B1 (HMGB1) is a nuclear protein that has been reported to contribute to tumor growth in humans. The present study identified a microRNA (miR/miRNA) that targets the 3' untranslated region (3'UTR) of the HMGB1 gene and assessed its effects on the proliferation of human cervical cancer cells and associated molecular mechanism. Western blotting was performed to determine HMGB1 levels in HeLa cells. TargetScan software was used to identify miRNA binding sites adjacent to the HMGB1. The viability of HeLa cells transfected with miR-142-3p mimics or inhibitors was determined using an MTT assay. The subcellular distribution (cytoplasmic or nuclear) of HMGB1 in HeLa cells was observed by western blotting. HMGB1 expression in HeLa and CaSKi cells was significantly higher compared with normal control cervical cells. TargetScan analysis indicated that miR-142-3p binds to the 3'UTR of HMGB1. Transfection with a miR-142-3p inhibitor increased cytoplasmic HMGB1 expression in HeLa cells, as shown by western blot analysis, while transfection with miR-142-3p mimics decreased the cytoplasmic expression of HMGB1 in HeLa cells. Therefore, miR-142-3p negatively regulated HMGB1 levels in cervical cancer cells. These findings indicated that miR-142-3p inhibited the proliferation of human cervical cancer cells, at least in part, by negatively regulating the cytoplasmic localization of HMGB1.

Ethyl Pyruvate Alleviates Pulmonary Hypertension through the Suppression of Pulmonary Artery Smooth Muscle Cell Proliferation via the High Mobility Group Protein B1/Receptor for Advanced Glycation End-Products Axis.

Purpose: Pulmonary arterial hypertension (PAH) is a formidable disease with no effective treatment at present. With the goal of developing potential therapies, we attempted to determine whether ethyl pyruvate (EP) could alleviate PAH and its mechanism.Methods: Pulmonary smooth muscle cells were cultured in conventional low-oxygen environments, and cellular proliferation was monitored after treatment with either EP or phosphate-balanced solution (PBS). Expression of high mobility group protein B1 (HMGB1) and receptor for advanced glycation end-products (RAGE) protein were detected by western blot. After hyperkinetic PAH rat models were treated with EP, hemodynamic data were collected. Right ventricular hypertrophy and pulmonary vascular remodeling were evaluated. Expression of HMGB1 and RAGE protein was also detected.Results: In vitro, proliferative activity increased in low-oxygen environments, but was inhibited by EP treatment. Furthermore, Western blotting showed the decreased expression of HMGB1 and RAGE protein after EP treatment. In vivo, pulmonary artery pressures were attenuated with EP. Right ventricular hypertrophy and pulmonary vascular remodeling were also reversed. Additionally, the expression levels of HMGB1 and RAGE were reduced in lung tissues.Conclusions: EP can alleviate PAH by suppressing the proliferation of pulmonary artery smooth muscle cells via inhibition of HMGB1/RAGE expression.

Hydrogen plays a protective role in intestinal injury of mice with severe sepsis by regulating the release of heme oxygenase-1 and high mobility group protein B1

To investigate the protective effect of the hydrogen (H2) inhalation on intestinal injury in severe sepsis mice and its relationship with nuclear factor E2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1)/high mobility group protein B1 (HMGB1) pathway. The 280 male wild-type (WT) and Nrf2 knockout (Nrf2-KO) ICR mice (6-8 weeks old, 20-25 g weight) were assigned into 8 groups randomly (WT and Nrf2-KO mice, 4 groups respectively, n = 35): Sham group, H2 inhalation group (Sham+H2 group), sepsis model group which was induced by cecal ligation and puncture (CLP group) and H2 treatment group (CLP+H2 group). In Sham group, except cecal ligation and perforation, the same surgery was used. The mice in Sham group and CLP group inhaled only air for 1 hour and 6 hours, respectively, while the mice in Sham+H2 group and CLP+H2 group inhaled 2% H2 for 60 minutes. After modeling, 20 mice randomly in each group were selected to measure 7-day natural survival rates; the remaining mice(15 mice) were killed in 24 hours after modeling, and the bacterial load was determined by collecting the peritoneal lavage fluid (PLF) of each mouse to measure the colony forming unit (CFU); and the intestinal tissue was taken after perfusion, and the intestinal tissue of 3 mice was used to extract protein for HO-1 and HMGB1 levels by using Western blotting. Intestinal tissue of 3 mice was extracted to prepare histomorphology sections for the identification of HO-1. Intestinal tissue of 3 mice was used to detect the expression of HMGB1 by immunofluorescence. The 7-day survival rate of WT and Nrf2-KO mice in CLP group was zero; in WT mice, the 7-day survival rate of CLP+H2 group was significantly higher than that of CLP group (45% vs. 0%, P < 0.05), while no difference of the 7-day survival rate in Nrf2-KO mice was noted between CLP and CLP+H2 groups (0% vs. 0%, P > 0.05). Compared with Sham group, the CFU in the PLF of both mice in CLP group increased significantly. Compared with CLP group, 2% H2 treatment could significantly improve the bacterial clearance rate of WT mice with severe sepsis (CFU, ×103: 34.7±6.3 vs. 74.2±8.1, P < 0.01), but had no significant effect on Nrf2-KO mice with severe sepsis (CFU, ×103: 73.3±7.5 vs. 75.8±8.6, P > 0.05). The results of Western blotting, immunohistochemical and immune-fluorescence showed that the expression of HO-1 and HMGB1 in the CLP group were higher than those in the Sham group. Compared with CLP group, the expression of HO-1 in WT mice of CLP+H2 group was significantly higher [HO-1 protein expression (HO-1/β-actin): 0.716±0.035 vs. 0.460±0.045, HO-1 positive expression (A value): 0.703±0.135 vs. 0.430±0.116, both P < 0.01], while the expression of HMGB1 was significantly lower [HMGB1 protein expression (HMGB1/β-actin): 0.052±0.038 vs. 0.082±0.008, HMGB1 positive expression (A value): 24.530±9.317 vs. 41.830±2.458, both P < 0.01]. In Nrf2-KO mice, compared with CLP group, there was no statistical difference in the above indexes in CLP+H2 group [HO-1 protein expression (HO-1/β-actin): 0.148±0.004 vs. 0.164±0.043, HO-1 positive expression (A value): 0.109±0.020 vs. 0.113±0.025, HMGB1 protein expression (HMGB1/β-actin): 0.109±0.020 vs. 0.113±0.025, HMGB1 positive expression (A value): 39.570±12.660 vs. 42.790±9.680, all P > 0.05]. H2 can inhibit the intestinal injury of mice with severe sepsis through Nrf2/HO-1/HMGB1 pathway, and Nrf2 plays an important role in the treatment of intestinal injury with H2.

Peripheral Cytokine Levels Differ by HPV Status and Change Treatment-Dependently in Patients with Head and Neck Squamous Cell Carcinoma.

Cytokines and immune mediators play an important role in the communication between immune cells guiding their response to infectious diseases or cancer. In this study, a comprehensive longitudinal analysis of serum cytokines and immune mediators in head and neck squamous cell carcinoma (HNSCC) patients was performed. In a prospective, non-interventional, longitudinal study, blood samples from 22 HNSCC patients were taken at defined time points (TP) before, during, and every 3 months after completion of (chemo)radio)therapy (CRT/RT) until 12 months after treatment. Serum concentrations of 17 cytokines/immune mediators and High-Mobility-Group-Protein B1 (HMGB1) were measured by fluorescent bead array and ELISA. Concentrations of sFas were significantly elevated during and after CRT/RT, whereas perforin levels were significantly decreased after CRT/RT. Levels of MIP-1β and Granzyme B differed significantly during CRT/RT by HPV status. Increased HMGB1 levels were observed at recurrence, accompanied by high levels of IL-4 and IL-10. The sFas increase and simultaneous perforin decrease may indicate an impaired immune cell function during adjuvant radiotherapy. Increased levels of pro-inflammatory cytokines in HPV+ compared to HPV− patients seem to reflect the elevated immunogenicity of HPV-positive tumors. High levels of HMGB1 and anti-inflammatory cytokines at recurrence may be interpreted as a sign of immune evasion.

Long Noncoding RNA HNF1A-AS1 Regulates Osteosarcoma Advancement Through Modulating the miR-32-5p/HMGB1 Axis.

Background: Osteosarcoma (OS) is a primary malignant tumor in children and adolescents. Long noncoding RNA HNF1A antisense RNA 1 (HNF1A-AS1) is connected with OS development. However, there are few reports on the role and mechanism of HNF1A-AS1 in OS. Materials and Methods: Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to assess the expression of HNF1A-AS1, miR-32-5p, and high-mobility group protein B1 (HMGB1). Western blot analysis was performed to detect the protein level of HMGB1. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), colony formation, transwell, or flow cytometer assays were applied to determine the proliferation, migration, invasion, and apoptosis of OS cells. The interaction between HNF1A-AS1 and miR-32-5p or HMGB1 was predicted by the starBase database and confirmed by dual-luciferase reporter assay. Enzyme-linked immunosorbent assay was employed to analyze levels of HMGB1 in the OS cell supernatant. Results: HNF1A-AS1 and HMGB1 were upregulated, while miR-32-5p was downregulated, in OS tissues and cells. Functionally, HNF1A-AS1 depletion induced apoptosis and impeded proliferation, migration, and invasion of OS cells. Interestingly, HNF1A-AS1 bound to miR-32-5p to regulate the expression of HMGB1. Furthermore, miR-32-5p knockdown overturned the effects of HNF1A-AS1 knockdown on apoptosis, proliferation, migration, and invasion of OS cells. In addition, the effects of HNF1A-AS1 silencing on the malignant behaviors of OS cells were reserved by HMGB1 overexpression. In addition, HNF1A-AS1 regulated the HMGB1 level in the OS cell supernatant through the miR-32-5p/HMGB1 axis. Conclusion: Downregulation of HNF1A-AS1 blocked OS progression through the miR-32-5p/HMGB1 axis, which provides a possible target and prognostic biomarker for treatment of OS.

High-mobility group protein B1: A predictive biomarker for hepatic encephalopathy after transjugular intrahepatic portosystemic shunt.

The aim of the present study was to investigate whether portal level of high-mobility group protein B1 (HMGB1) is associated with hepatic encephalopathy (HE) after transjugular intrahepatic portosystemic shunt (TIPS). We enrolled 127 consecutive patients who underwent TIPS and collected portal and peripheral blood samples in our department from December 2017 to May 2019. HMGB1 levels were determined using enzyme-linked immunosorbent assay kits. HMGB1 and other HE related parameters were estimated by competing risk analysis, receiver operating characteristic (ROC) analysis and Kaplan-Meier analysis. Patients with HE after TIPS were older (P = .019) and had higher portal HMGB1 level (P = .038) than those without. Univariate competing risk analysis: age (sHR 1.025, P=.026), hepatorenal syndrome (sHR 3.149, P=.010), model for end-of-stage liver disease (MELD) score (sHR 1.055, P=.024), prior HE (sHR 4.029, P=.0005), portal HMGB1 before TIPS (sHR 1.177, P = .001) reached statistical significance. Multivariate analysis: age (sHR 1.025, P=.037), MELD score (sHR 1.062, P=.011), prior HE (sHR 2.492, P=.030) and portal HMGB1 level before TIPS (sHR 1.217, P=.0002) were significantly different. ROC analyses and Kaplan-Meier curve showed portal HMGB1 level changes before and after TIPS (ΔHMGB1) had good predictive value in the cut-off 0.012 ng/mL (AUC=0.748, P < .001, Sensitivity=0.743, Specificity=0.655). Portal HMGB1 may be a therapeutic target for post-TIPS HE.

Inhibition of HMGB1 Overcomes Resistance to Radiation and Chemotherapy in Nasopharyngeal Carcinoma.

ObjectiveThis study aimed to investigate the effect of high mobility group protein B1 (HMGB1) on chemoresistance and radioresistance in nasopharyngeal carcinoma (NPC).Materials and MethodsHMGB1-knockout HK1 cell lines were generated using clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) system. Western blotting was used to evaluate the protein expression level of HMGB1. DNA repair efficiency of non-homologous end joining (NHEJ) and homologous recombination (HR) was monitored through NHEJ and HR reporter assay. Cellular protein–protein interaction between HMGB1 and NHEJ apparatus was determined by immunoprecipitation. Direct protein–protein interaction was examined by affinity capture assay with purified protein. Protein-DNA binding was evaluated by chromatin fractionation assay. Cell viability assay was employed to measure cell sensitivity to ionizing radiation (IR) or cisplatin.ResultsHMGB1-knockout NPC cells showed significant decrease in NHEJ efficiency. HMGB1 immunoprecipitated NHEJ key factors in NPC cells and promoted DNA-binding activity of Ku70. Mutational analysis revealed that serine 155 of Ku70 was required for its direct interaction with HMGB1. HMGB1 was highly expressed in radio- and chemoresistant NPC cells. Deficiency of HMGB1 sensitized wild-type (WT) and resistant NPC cells to IR and cisplatin. Glycyrrhizin, which is HMGB1 inhibitor, impaired DNA binding of HMGB1 and exhibited excellent synergy with IR and cisplatin.ConclusionHMGB1 promotes NHEJ via interaction with Ku70 resulting in resistance to IR and cisplatin. Inhibition of HMGB1 by glycyrrhizin is a potential therapeutic regimen to treat cisplatin and IR resistant NPC patients.

MiR-129-5p inhibits proliferation of gastric cancer cells through targeted inhibition on HMGB1 expression.

The aim of this study was to investigate the effects of micro ribonucleic acid (miR)-129-5p on the proliferation and apoptosis of gastric cancer cells via targeted repression on the expression of high mobility group protein B1 (HMGB1). Expression levels of miR-129-5p and HMGB1 in gastric cancer tissues (n=25) and adjacent normal tissues were measured via reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The regulatory effect of miR-129-5p on the proliferation of gastric cancer MGC-803 and SGC7901 cells was determined through Cell Counting Kit-8 (CCK-8) assay. Flow cytometry was employed to analyze the apoptosis rate of gastric cancer cells. To further discover the mechanism of miR-129-5p in regulating malignant behaviors of gastric cancer cells, the miRDB database was employed to predict the binding targets of miR-129-5p. Finally, binding sites of HMGB1 3'-untranslated region (3'-UTR) to miR-129-5p were discovered. Subsequently, HMGB1 wild-type or mutant 3'-UTR Luciferase reporter vectors were constructed, and transfected to MGC-803 and SGC7901 cells together with miR-129-5p or negative control miRNA. Next, Western blotting was adopted to measure the protein expression level of HMGB1 in MGC-803 and SGC7901 cells transfected with miR-129-5p or negative control miRNA, so as to investigate whether miR-129-5p affected HMGB1 protein expression. Additionally, to determine whether HMGB1 mediated the regulatory effect of miR-129-5p on the proliferation of gastric cancer cells, MGC-803 and SGC7901 cells were transfected with pcDNA-HMGB1 or pcDNA-vector, respectively. The expression level of HMGB1 was measured via RT-qPCR, and cell proliferation was determined by CCK-8 assay. The expression level of miR-129-5p in gastric cancer tissues was significantly lower than that in adjacent normal tissues (p<0.001). Meanwhile, the level of miR-129-5p was overtly lower in gastric cancer MGC-803 and SGC7901 cell lines than that in normal gastric mucosal epithelial GES-1 cells (p<0.001). These results indicated that miR-129-5p was lowly expressed in gastric cancer tissues and cell lines. Subsequent results demonstrated that the expression of HMGB1 increased remarkably in gastric cancer tissues compared with normal adjacent tissues (p<0.05). The proliferation ability of MGC-803 (p<0.001) and SGC7901 (p<0.01) cells with over-expressed miR-129-5p was remarkably weakened. Overexpression of miR-129-5p distinctly promoted the apoptosis rate of gastric cancer MGC-803 (p<0.01) and SGC7901 (p<0.001) cells. Moreover, miR-129-5p up-regulation significantly reduced the Luciferase activity of wild-type HMGB1 (p<0.001). However, no significant effect was observed on that of mutant HMGB1. The results suggested that overexpression of miR-129-5p significantly down-regulated the level of HMGB1 in gastric cancer cells. In addition, the messenger RNA (mRNA) level of HMGB1 in cells transfected with miR-129-5p also decreased significantly (p<0.001). HMGB1 overexpression overtly reversed the inhibitory effect of miR-129-5p on the proliferation of gastric cancer cells (p<0.05). All these results demonstrated that the miR-129-5p/HMGB1 axis played a key role in regulating the growth of gastric cancer cells. MiR-129-5p suppresses the progression of gastric cancer through targeted inhibition on the expression of HMGB1.

Decreased expression of prohibitin 1 mediates high-mobility group protein 1-induced pulmonary artery endothelial cell injury.

There is a significant increase of high-mobility group protein B1 (HMGB1) in plasma levels of patients with pulmonary hypertension, but the biological significance is still unclear. Anti-proliferative protein 1 (prohibitin 1, PHB1) is an important protein that maintains the homeostasis of vascular cells. This study aimed to investigate the effect of HMGB1 on pulmonary artery endothelial cells and the role of PHB1. In vivo experiment: A rat model of pulmonary hypertension induced by monocrotaline (MCT) was constructed. The right ventricular systolic pressure (RVSP), and the weight ratio of right ventricle to left ventricle plus ventricular septum were used to evaluate the success of model. ELISA was used to detect the level of HMGB1 in rat's plasma. Western blotting was used to detect the level of PHB1 in rat's lung tissues. CD31 immunofluorescence was used to detect the integrity of pulmonary vascular endothelium. In vitro experiments: Pulmonary artery endothelial cell (PAEC) was incubated with HMGB1 to observe the effect of HMGB1 on PAEC injury. Overexpression and knockdown of PHB1 were conducted, and the role of PHB1 was investigated by detecting the levels of reative oxygen species and cytochrome c (cyto-c), and the activation of caspase-3. Compared with the control group, the level of HMGB1 in the plasma of rats with pulmonary hypertension was significantly increased (P<0.05), and the expression of PHB1 in the lung tissue was decreased accompanied with endothelial dysfunction (P<0.05); HMGB1 incubation damaged the pulmonary artery endothelium and down-regulated PHB1 expression (P<0.05), while overexpression of PHB1 reduced the PAEC damage and oxidative stress induced by HMGB1 (P<0.05). Meanwhile, PHB1 reduced HMGB1-induced cyto-c expression and caspase-3 cleavage by inhibiting oxidative stress (P<0.05). The down-regulation of PHB1 expression mediates HMGB1-induced PAEC injury, which is related to the induction of oxidative stress, the increase of cyto-c release, and the promotion of caspase-3 cleavage.