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1.
World J Exp Med ; 14(2): 90374, 2024 Jun 20.
Article in English | MEDLINE | ID: mdl-38948415

ABSTRACT

BACKGROUND: ATP sensitive K+ (KATP) channels are ubiquitously distributed in various of cells and tissues, including the liver. They play a role in the pathogenesis of myocardial and liver ischemia. AIM: To evaluate the radiation-induced changes in the expression of KATP channel subunits in the mouse liver to understand the potential role of KATP channels in radiation injury. METHODS: Adult C57BL/6 mice were randomly exposed to γ-rays at 0 Gy (control, n = 2), 0.2 Gy (n = 6), 1 Gy (n = 6), or 5 Gy (n = 6). The livers were removed 3 and 24 h after radiation exposure. Hematoxylin and eosin staining was used for morphological observation; immunohistochemical staining was applied to determine the expression of KATP channel subunits in the liver tissue. RESULTS: Compared with the control group, the livers exposed to 0.2 Gy γ-ray showed an initial increase in the expression of Kir6.1 at 3 h, followed by recovery at 24 h after exposure. Exposure to a high dose of 5.0 Gy resulted in decreased expression of Kir6.1 and increased expression of SUR2B at 24 h. However, the expression of Kir6.2, SUR1, or SUR2A had no remarkable changes at 3 and 24 h after exposure to any of these doses. CONCLUSION: The expression levels of Kir6.1 and SUR2B in mouse liver changed differently in response to different radiation doses, suggesting a potential role for them in radiation-induced liver injury.

3.
Br J Pharmacol ; 2024 Jul 01.
Article in English | MEDLINE | ID: mdl-38952084

ABSTRACT

BACKGROUND AND PURPOSE: The gut hormone glucose-dependent insulinotropic polypeptide (GIP) signals via the GIP receptor (GIPR), resulting in postprandial potentiation of glucose-stimulated insulin secretion. The translation of results from rodent studies to human studies has been challenged by the unexpected effects of GIPR-targeting compounds. We, therefore, investigated the variation between species, focusing on GIPR desensitization and the role of the receptor C-terminus. EXPERIMENTAL APPROACH: The GIPR from humans, mice, rats, pigs, dogs and cats was studied in vitro for cognate ligand affinity, G protein activation (cAMP accumulation), recruitment of beta-arrestin and internalization. Variants of the mouse, rat and human GIPRs with swapped C-terminal tails were studied in parallel. KEY RESULTS: The human GIPR is more prone to internalization than rodent GIPRs. Despite similar agonist affinities and potencies for Gαs activation, especially, the mouse GIPR shows reduced receptor desensitization, internalization and beta-arrestin recruitment. Using an enzyme-stabilized, long-acting GIP analogue, the species differences were even more pronounced. 'Tail-swapped' human, rat and mouse GIPRs were all fully functional in their Gαs coupling, and the mouse GIPR regained internalization and beta-arrestin 2 recruitment properties with the human tail. The human GIPR lost the ability to recruit beta-arrestin 2 when its own C-terminus was replaced by the rat or mouse tail. CONCLUSIONS AND IMPLICATIONS: Desensitization of the human GIPR is dependent on the C-terminal tail. The species-dependent functionality of the C-terminal tail and the different species-dependent internalization patterns, especially between human and mouse GIPRs, are important factors influencing the preclinical evaluation of GIPR-targeting therapeutic compounds.

4.
Zhongguo Xue Xi Chong Bing Fang Zhi Za Zhi ; 36(3): 286-293, 2024 Jun 17.
Article in Chinese | MEDLINE | ID: mdl-38952315

ABSTRACT

OBJECTIVE: To investigate the involvement of the high mobility group box protein B1 (HMGB1)-Toll-like receptor 2 (TLR2)/TLR4-nuclear factor κB (NF-κB) pathway in the intestinal mucosal injury induced by Cryptosporidium parvum infection, and to examine the effect of oxymatrine (OMT) on C. parvum infection in mice. METHODS: Forty SPF 4-week-old BALB/c mice were randomly divided into four groups, including the control group, infection group, glycyrrhizin (GA) group and OMT group. Each mouse was orally administered with 1 × 105 C. parvum oocysts one week in the infection, GA and OMT groups following dexamethasone-induced immunosuppression to model C. parvum intestinal infections in mice. Upon successful modeling, mice in the GA group were intraperitoneally injected with GA at a daily dose of 25.9 mL/kg for successive two weeks, and animals in the OMT group were orally administered OMT at a daily dose of 50 mg/kg for successive two weeks, while mice in the control group were given normal food and water. All mice were sacrificed two weeks post-treatment, and proximal jejunal tissues were sampled. The pathological changes of mouse intestinal mucosal specimens were observed using hematoxylin-eosin (HE) staining, and the mouse intestinal villous height, intestinal crypt depth and the ratio of intestinal villous height to intestinal crypt depth were measured. The occludin and zonula occludens protein 1 (ZO1) expression was determined in mouse intestinal epithelial cells using immunohistochemistry, and the relative expression of HMGB1, TLR2, TLR4, myeloid differentiation primary response gene 88 (MyD88) and NF-κB p65 mRNA was quantified in mouse jejunal tissues using quantitative real-time PCR (qPCR) assay. RESULTS: HE staining showed that the mouse intestinal villi were obviously atrophic, shortened, and detached, and the submucosal layer of the mouse intestine was edematous in the infection group as compared with the control group, while the mouse intestinal villi tended to be structurally intact and neatly arranged in the GA and OMT groups. There were significant differences among the four groups in terms of the mouse intestinal villous height (F = 6.207, P = 0.000 5), intestinal crypt depth (F = 6.903, P = 0.000 3) and the ratio of intestinal villous height to intestinal crypt depth (F = 37.190, P < 0.000 1). The mouse intestinal villous height was lower in the infection group than in the control group [(321.9 ± 41.1) µm vs. (399.5 ± 30.9) µm; t = 4.178, P < 0.01] and the GA group [(321.9 ± 41.1) µm vs. (383.7 ± 42.7) µm; t = 3.130, P < 0.01], and the mouse intestinal crypt depth was greater in the infection group [(185.0 ± 35.9) µm] than in the control group [(128.4 ± 23.6) µm] (t = 3.877, P < 0.01) and GA group [(143.3 ± 24.7) µm] (t = 2.710, P < 0.05). The mouse intestinal villous height was greater in the OMT group [(375.3 ± 22.9) µm] than in the infection group (t = 3.888, P < 0.01), and there was no significant difference in mouse intestinal villous height between the OMT group and the control group (t = 1.989, P > 0.05). The mouse intestinal crypt depth was significantly lower in the OMT group [(121.5 ± 27.3) µm] than in the infection group (t = 4.133, P < 0.01), and there was no significant difference in mouse intestinal crypt depth between the OMT group and the control group (t = 0.575, P > 0.05). The ratio of the mouse intestinal villous height to intestinal crypt depth was significantly lower in the infection group (1.8 ± 0.2) than in the control group (3.1 ± 0.3) (t = 10.540, P < 0.01) and the GA group (2.7 ± 0.3) (t = 7.370, P < 0.01), and the ratio of the mouse intestinal villous height to intestinal crypt depth was significantly higher in the OMT group (3.1 ± 0.2) than in the infection group (t = 15.020, P < 0.01); however, there was no significant difference in the ratio of the mouse intestinal villous height to intestinal crypt depth between the OMT group and the control group (t = 0.404, P > 0.05). Immunohistochemical staining showed significant differences among the four groups in terms of occludin (F = 28.031, P < 0.000 1) and ZO1 expression (F = 14.122, P < 0.000 1) in mouse intestinal epithelial cells. The proportion of positive occluding expression was significantly lower in mouse intestinal epithelial cells in the infection group than in the control group [(14.3 ± 4.5)% vs. (28.3 ± 0.5)%; t = 3.810, P < 0.01], and the proportions of positive occluding expression were significantly higher in mouse intestinal epithelial cells in the GA group [(30.3 ± 1.3)%] and OMT group [(25.8 ± 1.5)%] than in the infection group (t = 7.620 and 5.391, both P values < 0.01); however, there was no significant differences in the proportion of positive occluding expression in mouse intestinal epithelial cells between the GA or OMT groups and the control group (t = 1.791 and 2.033, both P values > 0.05). The proportion of positive ZO1 expression was significantly lower in mouse intestinal epithelial cells in the infection group than in the control group [(14.4 ± 1.8)% vs. (24.2 ± 2.8)%; t = 4.485, P < 0.01], and the proportions of positive ZO1 expression were significantly higher in mouse intestinal epithelial cells in the GA group [(24.1 ± 2.3)%] (t = 5.159, P < 0.01) and OMT group than in the infection group [(22.5 ± 1.9)%] (t = 4.441, P < 0.05); however, there were no significant differences in the proportion of positive ZO1 expression in mouse intestinal epithelial cells between the GA or OMT groups and the control group (t = 0.037 and 0.742, both P values > 0.05). qPCR assay showed significant differences among the four groups in terms of HMGB1 (F = 21.980, P < 0.000 1), TLR2 (F = 20.630, P < 0.000 1), TLR4 (F = 17.000, P = 0.000 6), MyD88 (F = 8.907, P = 0.000 5) and NF-κB p65 mRNA expression in mouse jejunal tissues (F = 8.889, P = 0.000 7). The relative expression of HMGB1 [(5.97 ± 1.07) vs. (1.05 ± 0.07); t = 6.482, P < 0.05] 、TLR2 [(5.92 ± 1.29) vs. (1.10 ± 0.14); t = 5.272, P < 0.05] 、TLR4 [(5.96 ± 1.50) vs. (1.02 ± 0.03); t = 4.644, P < 0.05] 、MyD88 [(3.00 ± 1.26) vs. (1.02 ± 0.05); t = 2.734, P < 0.05] and NF-κB p65 mRNA [(2.33 ± 0.72) vs. (1.04 ± 0.06); t = 2.665, P < 0.05] was all significantly higher in mouse jejunal tissues in the infection group than in the control group. A significant reduction was detected in the relative expression of HMGB1 (0.63 ± 0.01), TLR2 (0.42 ± 0.10), TLR4 (0.35 ± 0.07), MyD88 (0.70 ± 0.11) and NF-κB p65 mRNA (0.75 ± 0.01) in mouse jejunal tissues in the GA group relative to the control group (t = 8.629, 5.830, 11.500, 4.729 and 6.898, all P values < 0.05), and the relative expression of HMGB1, TLR2, TLR4, MyD88 and NF-κB p65 mRNA significantly reduced in mouse jejunal tissues in the GA group as compared to the infection group (t = 7.052, 6.035, 4.084, 3.165 and 3.274, all P values < 0.05). In addition, the relative expression of HMGB1 (1.14 ± 0.60), TLR2 (1.00 ± 0.24), TLR4 (1.14 ± 0.07), MyD88 (0.96 ± 0.25) and NF-κ B p65 mRNA (1.12 ± 0.17) was significantly lower in mouse jejunal tissues in the OMT group than in the infection group (t = 7.059, 5.320, 3.510, 3.466 and 3.273, all P values < 0.05); however, there were no significant differences between the OMT and control groups in terms of relative expression of HMGB1, TLR2, TLR4, MyD88 or NF-κB p65 mRNA in mouse jejunal tissues (t = 0.239, 0.518, 1.887, 0.427 and 0.641, all P values > 0.05). CONCLUSIONS: C. parvum infection causes intestinal inflammatory responses and destruction of intestinal mucosal barrier through up-regulating of the HMGB1-TLR2/TLR4-NF-κB pathway. OMT may suppress the intestinal inflammation and repair the intestinal mucosal barrier through inhibiting the activity of the HMGB1-TLR2/TLR4-NF-κB pathway.


Subject(s)
Alkaloids , Cryptosporidiosis , Cryptosporidium parvum , HMGB1 Protein , Mice, Inbred BALB C , NF-kappa B , Quinolizines , Toll-Like Receptor 2 , Toll-Like Receptor 4 , Animals , Cryptosporidiosis/drug therapy , Cryptosporidiosis/parasitology , Quinolizines/pharmacology , Cryptosporidium parvum/drug effects , Cryptosporidium parvum/physiology , Toll-Like Receptor 4/genetics , Toll-Like Receptor 4/metabolism , Mice , Toll-Like Receptor 2/metabolism , Toll-Like Receptor 2/genetics , NF-kappa B/metabolism , NF-kappa B/genetics , Alkaloids/pharmacology , Alkaloids/administration & dosage , HMGB1 Protein/metabolism , HMGB1 Protein/genetics , Signal Transduction/drug effects , Male , Intestinal Mucosa/drug effects , Intestinal Mucosa/parasitology , Intestinal Mucosa/metabolism , Matrines
5.
Zhongguo Xue Xi Chong Bing Fang Zhi Za Zhi ; 36(3): 304-309, 2024 Jun 07.
Article in Chinese | MEDLINE | ID: mdl-38952318

ABSTRACT

OBJECTIVE: To investigate the development and dynamic changes of cysts in the brain of mice following infection with different forms of Toxoplasma gondii, so as to provide insights into for toxoplasmosis prevention and control. METHODS: ICR mice at ages of 6 to 8 weeks, each weighing 20 to 25 g, were intraperitoneally injected with tachyzoites of the T. gondii PRU strain at a dose of 1 × 105 tachyzoites per mouse, orally administered with cysts at a dose of 20 oocysts per mouse or oocysts at a dose of 200 oocysts per mouse for modeling chronic T. gondii infection in mice, and the clinical symptoms and survival of mice were observed post-infection. Mice were orally infected with T. gondii cysts at doses of 10 (low-dose group), 20 (medium-dose group), 40 cysts per mouse (high-dose group), and the effect of different doses of T. gondii infections on the number of cysts was examined in the mouse brain. Mice were orally administered with T. gondii cysts at a dose of 20 cysts per mouse, and grouped according to gender (female and male) and time points of infections (20, 30, 60, 90, 120, 150, 180 days post-infection), and the effects of gender and time points of infections on the number of cysts was examined in the mouse brain. In addition, mice were divided into the tachyzoite group (Group T), the first-generation cyst group (Group C1), the second-generation cyst group (Group C2), the third-generation cyst (Group C3) and the fourth-generation cyst group (Group C4). Mice in the Group T were intraperitoneally injected with T. gondii tachyzoites at a dose of 1 × 105 tachyzoites per mouse, and the cysts were collected from the mouse brain tissues 30 days post-infection, while mice in the Group C1 were orally infected with the collected cysts at a dose of 30 cysts per mouse. Continuous passage was performed by oral administration with cysts produced by the previous generation in mice, and the effect of continuous passage on the number of cysts was examined in the mouse brain. RESULTS: Following infection with T. gondii tachyzoites, cysts and oocysts in mice, obvious clinical symptoms were observed on days 6 to 13 and mice frequently died on days 7 to 12. The survival rates of mice were 67.0%, 87.0% and 53.0%, and the mean numbers of cysts were (516.0 ± 257.2), (1 203.0 ± 502.0) and (581.0 ± 183.1) in the mouse brain (F = 11.94, P < 0.01) on day 30 post-infection with T. gondii tachyzoites, cysts and oocysts, respectively, and the numbers of cysts in the brain tissues were significantly lower in mice infected with T. gondii tachyzoites and oocysts than in those infected with cysts (all P values < 0.01). The survival rates of mice were 87.0%, 87.0% and 60.0%, and the mean numbers of cysts were (953.0 ± 355.5), (1 084.0 ± 474.3) and (1 113.0 ± 546.0) in the mouse brain in the low-, medium- and high-dose groups on day 30 post-infection, respectively (F = 0.42, P > 0.05). The survival rates of male and female mice were 73.0% and 80.0%, and the mean numbers of cysts were (946.4 ± 411.4) and (932.1 ± 322.4) in the brain tissues of male and female mice, respectively (F = 1.63, P > 0.05). Following continuous passage, the mean numbers of cysts were (516.0 ± 257.2), (1 203.0 ± 502.0), (896.8 ± 332.3), (782.5 ± 423.9) and (829.2 ± 306.0) in the brain tissues of mice in the T, C1, C2, C3 and C4 groups, respectively (F = 4.82, P < 0.01), and the number of cysts was higher in the mouse brain in Group 1 than in Group T (P < 0.01). Following oral administration of 20 T. gondii cysts in mice, cysts were found in the moues brain for the first time on day 20 post-infection, and the number of cysts gradually increased over time, peaked on days 30 and 90 post-infection and then gradually decreased; however, the cysts were still found in the mouse brain on day 180 post-infection. CONCLUSIONS: There is a higher possibility of developing chronic T. gondii infection in mice following infection with cysts than with oocysts or tachyzoites and the most severe chronic infection is seen following infection with cysts. The number of cysts does not correlate with the severity of chronic T. gondii infection, and the number of cysts peaks in the mouse brain on days 30 and 90 post-infection.


Subject(s)
Brain , Mice, Inbred ICR , Toxoplasma , Toxoplasmosis, Animal , Animals , Mice , Female , Male , Brain/parasitology , Chronic Disease , Toxoplasmosis, Animal/parasitology , Toxoplasma/physiology , Toxoplasmosis/parasitology , Disease Models, Animal
6.
Elife ; 132024 Jul 01.
Article in English | MEDLINE | ID: mdl-38949655

ABSTRACT

Secreted chemokines form concentration gradients in target tissues to control migratory directions and patterns of immune cells in response to inflammatory stimulation; however, how the gradients are formed is much debated. Heparan sulfate (HS) binds to chemokines and modulates their activities. In this study, we investigated the roles of HS in the gradient formation and chemoattractant activity of CCL5 that is known to bind to HS. CCL5 and heparin underwent liquid-liquid phase separation and formed gradient, which was confirmed using CCL5 immobilized on heparin-beads. The biological implication of HS in CCL5 gradient formation was established in CHO-K1 (wild-type) and CHO-677 (lacking HS) cells by Transwell assay. The effect of HS on CCL5 chemoattractant activity was further proved by Transwell assay of human peripheral blood cells. Finally, peritoneal injection of the chemokines into mice showed reduced recruitment of inflammatory cells either by mutant CCL5 (lacking heparin-binding sequence) or by addition of heparin to wild-type CCL5. Our experimental data propose that co-phase separation of CCL5 with HS establishes a specific chemokine concentration gradient to trigger directional cell migration. The results warrant further investigation on other heparin-binding chemokines and allows for a more elaborate insight into disease process and new treatment strategies.


Subject(s)
Chemokine CCL5 , Chemotaxis , Cricetulus , Heparitin Sulfate , Chemokine CCL5/metabolism , Chemokine CCL5/genetics , Animals , Heparitin Sulfate/metabolism , Humans , CHO Cells , Mice , Heparin/metabolism , Heparin/pharmacology , Phase Separation
7.
Front Immunol ; 15: 1425938, 2024.
Article in English | MEDLINE | ID: mdl-38953020

ABSTRACT

Introduction: P2X receptors are a family of homo- and heterotrimeric cation channels gated by extracellular ATP. The P2X4 and P2X7 subunits show overlapping expression patterns and have been involved in similar physiological processes, such as pain and inflammation as well as various immune cell functions. While formation of P2X2/P2X3 heterotrimers produces a distinct pharmacological phenotype and has been well established, functional identification of a P2X4/P2X7 heteromer has been difficult and evidence for and against a physical association has been found. Most of this evidence stems, however, from in vitro model systems. Methods: Here, we used a P2X7-EGFP BAC transgenic mouse model as well as P2X4 and P2X7 knock-out mice to re-investigate a P2X4-P2X7 interaction in mouse lung by biochemical and immunohistochemical experiments as well as quantitative expression analysis. Results: No detectable amounts of P2X4 could be co-purified from mouse lung via P2X7-EGFP. In agreement with these findings, immuno-histochemical analysis using a P2X7-specific nanobody revealed only limited overlap in the cellular and subcellular localizations of P2X4 and P2X7 in both the native lung tissue and primary cells. Comparison of P2X4 and P2X7 transcript and protein levels in the respective gene-deficient and wild type mice showed no mutual interrelation between their expression levels in whole lungs. However, a significantly reduced P2rx7 expression was found in alveolar macrophages of P2rx4 -/- mice. Discussion: In summary, our detailed analysis of the cellular and subcellular P2X4 and P2X7 localization and expression does not support a physiologically relevant direct association of P2X4 and P2X7 subunits or receptors in vivo.


Subject(s)
Lung , Mice, Knockout , Mice, Transgenic , Receptors, Purinergic P2X4 , Receptors, Purinergic P2X7 , Animals , Receptors, Purinergic P2X4/metabolism , Receptors, Purinergic P2X4/genetics , Receptors, Purinergic P2X7/genetics , Receptors, Purinergic P2X7/metabolism , Mice , Lung/metabolism , Lung/immunology , Mice, Inbred C57BL , Protein Binding
8.
J Cell Commun Signal ; 18(2): e12033, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38946724

ABSTRACT

Liver fibrosis is a persistent damage repair response triggered by various injury factors, which leads to an abnormal accumulation of extracellular matrix within liver tissue samples. The current clinical treatment of liver fibrosis is currently ineffective; therefore, elucidating the mechanism of liver fibrogenesis is of significant importance. Herein, the function and related mechanisms of lncRNA Snhg12 within hepatic fibrosis were investigated. Snhg12 expression was shown to be increased in mouse hepatic fibrotic tissue samples, and Snhg12 knockdown suppressed hepatic pathological injury and down-regulated the expression levels of fibrosis-associated proteins. Mechanistically, Snhg12 played a role in the early activation of mouse hepatic stellate cells (mHSCs) based on bioinformatics analysis, and Snhg12 was positively correlated with Igfbp3 expression. Further experimental results demonstrated that Snhg12 knockdown impeded mHSCs proliferation and activation and also downregulated the protein expression of Igfbp3. Snhg12 could interact with IGFBP3 and boost its protein stability, and overexpression of Igfbp3 partially reversed the inhibition of mHSCsproliferation and activation by the knockdown of Snhg12. In conclusion, LncRNA Snhg12 mediates liver fibrosis by targeting IGFBP3 and promoting its protein stability, thereby promoting mHSC proliferation and activation. Snhg12 has been identified as an underlying target for treating liver fibrosis.

9.
Front Cell Dev Biol ; 12: 1363541, 2024.
Article in English | MEDLINE | ID: mdl-38946797

ABSTRACT

Introduction: Duchenne muscular dystrophy (DMD) is a genetic disorder caused by mutations in the dystrophin-encoding gene that leads to muscle necrosis and degeneration with chronic inflammation during growth, resulting in progressive generalized weakness of the skeletal and cardiac muscles. We previously demonstrated the therapeutic effects of systemic administration of dental pulp mesenchymal stromal cells (DPSCs) in a DMD animal model. We showed preservation of long-term muscle function and slowing of disease progression. However, little is known regarding the effects of cell therapy on the metabolic abnormalities in DMD. Therefore, here, we aimed to investigate the mechanisms underlying the immunosuppressive effects of DPSCs and their influence on DMD metabolism. Methods: A comprehensive metabolomics-based approach was employed, and an ingenuity pathway analysis was performed to identify dystrophy-specific metabolomic impairments in the mdx mice to assess the therapeutic response to our established systemic DPSC-mediated cell therapy approach. Results and Discussion: We identified DMD-specific impairments in metabolites and their responses to systemic DPSC treatment. Our results demonstrate the feasibility of the metabolomics-based approach and provide insights into the therapeutic effects of DPSCs in DMD. Our findings could help to identify molecular marker targets for therapeutic intervention and predict long-term therapeutic efficacy.

10.
Cancer Innov ; 3(3): e118, 2024 Jun.
Article in English | MEDLINE | ID: mdl-38947755

ABSTRACT

Background: Cancer-targeted T-cell receptor T (TCR-T) cells hold promise in treating cancers such as hematological malignancies and breast cancers. However, approaches to obtain cancer-reactive TCR-T cells have been unsuccessful. Methods: Here, we developed a novel strategy to screen for cancer-targeted TCR-T cells using a special humanized mouse model with person-specific immune fingerprints. Rare steady-state circulating hematopoietic stem and progenitor cells were expanded via three-dimensional culture of steady-state peripheral blood mononuclear cells, and then the expanded cells were applied to establish humanized mice. The human immune system was evaluated according to the kinetics of dendritic cells, monocytes, T-cell subsets, and cytokines. To fully stimulate the immune response and to obtain B-cell precursor NAML-6- and triple-negative breast cancer MDA-MB-231-targeted TCR-T cells, we used the inactivated cells above to treat humanized mice twice a day every 7 days. Then, human T cells were processed for TCR ß-chain (TRB) sequencing analysis. After the repertoires had been constructed, features such as the fraction, diversity, and immune signature were investigated. Results: The results demonstrated an increase in diversity and clonality of T cells after treatment. The preferential usage and features of TRBV, TRBJ, and the V-J combination were also changed. The stress also induced highly clonal expansion. Tumor burden and survival analysis demonstrated that stress induction could significantly inhibit the growth of subsequently transfused live tumor cells and prolong the survival of the humanized mice. Conclusions: We constructed a personalized humanized mouse model to screen cancer-targeted TCR-T pools. Our platform provides an effective source of cancer-targeted TCR-T cells and allows for the design of patient-specific engineered T cells. It therefore has the potential to greatly benefit cancer treatment.

12.
Aging Cell ; : e14259, 2024 Jul 03.
Article in English | MEDLINE | ID: mdl-38961628

ABSTRACT

Hutchinson-Gilford progeria syndrome (HGPS) is a rare premature aging disorder resulting from de novo mutations in the lamin A gene. Children with HGPS typically pass away in their teenage years due to cardiovascular diseases such as atherosclerosis, myocardial infarction, heart failure, and stroke. In this study, we characterized the G608G HGPS mouse model and explored cardiac and skeletal muscle function, along with senescence-associated phenotypes in fibroblasts. Homozygous G608G HGPS mice exhibited cardiac dysfunction, including decreased cardiac output and stroke volume, and impaired left ventricle relaxation. Additionally, skeletal muscle exhibited decreased isometric tetanic torque, muscle atrophy, and increased fibrosis. HGPS fibroblasts showed nuclear abnormalities, decreased proliferation, and increased expression of senescence markers. These findings provide insights into the pathophysiology of the G608G HGPS mouse model and inform potential therapeutic strategies for HGPS.

13.
Sci Rep ; 14(1): 15066, 2024 07 02.
Article in English | MEDLINE | ID: mdl-38956113

ABSTRACT

Living cells have spontaneous ultraweak photon emission derived from metabolic reactions associated with physiological conditions. The ORCA-Quest CMOS camera (Hamamatsu Photonics, Japan) is a highly sensitive and essential tool for photon detection; its use with a microscope incubator (Olympus) enables the detection of photons emitted by embryos with the exclusion of harmful visible light. With the application of the second law of thermodynamics, the low-entropy energy absorbed and used by embryos can be distinguished from the higher-entropy energy released and detectable in their environment. To evaluate higher-entropy energy data from embryos, we developed a unique algorithm for the calculation of the entropy-weighted spectral fractal dimension, which demonstrates the self-similar structure of the energy (photons) released by embryos. Analyses based on this structure enabled the distinction of living and degenerated mouse embryos, and of frozen and fresh embryos and the background. This novel detection of ultra-weak photon emission from mouse embryos can provide the basis for the development of a photon emission embryo control system. The ultraweak photon emission fingerprints of embryos may be used for the selection of viable specimens in an ideal dark environment.


Subject(s)
Algorithms , Embryo, Mammalian , Photons , Animals , Mice , Female
14.
BMC Cancer ; 24(1): 792, 2024 Jul 03.
Article in English | MEDLINE | ID: mdl-38956496

ABSTRACT

The in vivo functions of SerpinB2 in tumor cells and tumor-associated macrophages (TAMs) during breast cancer development and metastasis remain elusive. SerpinB2-deficient MMTV-PyMT mice (PyMTSB2-/-) were previously produced to explore the biological roles of SerpinB2 in breast cancer. Compared with MMTV-PyMT wild-type (PyMTWT) mice, PyMTSB2-/- mice showed delayed tumor progression and reduced CK8 + tumor cell dissemination to lymph nodes. RNA-Seq data revealed significantly enriched genes associated with inflammatory responses, especially upregulated M1 and downregulated M2 macrophage marker genes in PyMTSB2-/- tumors. Decreased CD206+M2 and increased NOS2+M1 markers were detected in the primary tumors and metastatic lymph nodes of PyMTSB2-/- mice. In an in vitro study, SerpinB2 knockdown decreased the sphere formation and migration of MDA-MB-231 cells and suppressed protumorigenic M2 polarization of RAW264.7 cells. The combination of low SerpinB2, high NOS2, and low CD206 expression was favorable for survival in patients with breast cancer, as assessed in the BreastMark dataset. Our study demonstrates that SerpinB2 deficiency delays mammary tumor development and metastasis in PyMTWT mice, along with reduced sphere formation and migration abilities of tumor cells and decreased macrophage protumorigenic polarization.


Subject(s)
Breast Neoplasms , Plasminogen Activator Inhibitor 2 , Animals , Mice , Female , Plasminogen Activator Inhibitor 2/genetics , Plasminogen Activator Inhibitor 2/metabolism , Plasminogen Activator Inhibitor 2/deficiency , Humans , Breast Neoplasms/pathology , Breast Neoplasms/genetics , Breast Neoplasms/metabolism , Macrophages/metabolism , Tumor-Associated Macrophages/metabolism , Cell Line, Tumor , Mice, Knockout , RAW 264.7 Cells , Mammary Neoplasms, Experimental/pathology , Mammary Neoplasms, Experimental/genetics , Nitric Oxide Synthase Type II/metabolism , Nitric Oxide Synthase Type II/genetics , Cell Movement/genetics
15.
Cell Biosci ; 14(1): 88, 2024 Jul 02.
Article in English | MEDLINE | ID: mdl-38956702

ABSTRACT

This study investigates NADPH oxidase 4 (NOX4) involvement in iron-mediated astrocyte cell death in Alzheimer's Disease (AD) using single-cell sequencing data and transcriptomes. We analyzed AD single-cell RNA sequencing data, identified astrocyte marker genes, and explored biological processes in astrocytes. We integrated AD-related chip data with ferroptosis-related genes, highlighting NOX4. We validated NOX4's role in ferroptosis and AD in vitro and in vivo. Astrocyte marker genes were enriched in AD, emphasizing their role. NOX4 emerged as a crucial player in astrocytic ferroptosis in AD. Silencing NOX4 mitigated ferroptosis, improved cognition, reduced Aß and p-Tau levels, and alleviated mitochondrial abnormalities. NOX4 promotes astrocytic ferroptosis, underscoring its significance in AD progression.

16.
Redox Biol ; 75: 103261, 2024 Jun 28.
Article in English | MEDLINE | ID: mdl-38963974

ABSTRACT

Squamous cell carcinomas (SCCs), including lung, head & neck, bladder, and skin SCCs often display constitutive activation of the KEAP1-NRF2 pathway. Constitutive activation is achieved through multiple mechanisms, including activating mutations in NFE2L2 (NRF2). To determine the functional consequences of Nrf2 activation on skin SCC development, we assessed the effects of mutant Nrf2E79Q expression, one of the most common activating mutations in human SCCs, on tumor promotion and progression in the mouse skin multistage carcinogenesis model using a DMBA-initiation/TPA-promotion protocol where the Hras A->T mutation (Q61L) is the canonical driver mutation. Nrf2E79Q expression was temporally and conditionally activated in the epidermis at two stages of tumor development: 1) after DMBA initiation in the epidermis but before cutaneous tumor development and 2) in pre-existing DMBA-initiated/TPA-promoted squamous papillomas. Expression of Nrf2E79Q in the epidermis after DMBA initiation but before tumor occurrence inhibited the development/promotion of 70% of squamous papillomas. However, the remaining papillomas often displayed non-canonical Hras and Kras mutations and enhanced progression to SCCs compared to control mice expressing wildtype Nrf2. Nrf2E79Q expression in pre-existing tumors caused rapid regression of 60% of papillomas. The remaining papillomas displayed the expected canonical Hras A->T mutation (Q61L) and enhanced progression to SCCs. These results demonstrate that mutant Nrf2E79Q enhances the promotion and progression of a subset of skin tumors and alters the frequency and diversity of oncogenic Ras mutations when expressed early after initiation.

17.
Elife ; 132024 Jul 04.
Article in English | MEDLINE | ID: mdl-38963781

ABSTRACT

Reports indicate that an interaction between TRPV4 and anoctamin 1 (ANO1) could be widely involved in water efflux of exocrine glands, suggesting that the interaction could play a role in perspiration. In secretory cells of sweat glands present in mouse foot pads, TRPV4 clearly colocalized with cytokeratin 8, ANO1, and aquaporin-5 (AQP5). Mouse sweat glands showed TRPV4-dependent cytosolic Ca2+ increases that were inhibited by menthol. Acetylcholine-stimulated sweating in foot pads was temperature-dependent in wild-type, but not in TRPV4-deficient mice and was inhibited by menthol both in wild-type and TRPM8KO mice. The basal sweating without acetylcholine stimulation was inhibited by an ANO1 inhibitor. Sweating could be important for maintaining friction forces in mouse foot pads, and this possibility is supported by the finding that wild-type mice climbed up a slippery slope more easily than TRPV4-deficient mice. Furthermore, TRPV4 expression was significantly higher in controls and normohidrotic skin from patients with acquired idiopathic generalized anhidrosis (AIGA) compared to anhidrotic skin from patients with AIGA. Collectively, TRPV4 is likely involved in temperature-dependent perspiration via interactions with ANO1, and TRPV4 itself or the TRPV4/ANO 1 complex would be targeted to develop agents that regulate perspiration.


Stress, spicy foods and elevated temperatures can all trigger specialized gland cells to move water to the skin ­ in other words, they can make us sweat. This process is one of the most important ways by which our bodies regulate their temperature and avoid life-threatening conditions such as heatstroke. Disorders in which this function is impaired, such as AIGA (acquired idiopathic generalized anhidrosis), pose significant health risks. Finding treatments for sweat-related diseases requires a detailed understanding of the molecular mechanisms behind sweating, which has yet to be achieved. Recent research has highlighted the role of two ion channels, TRPV4 and ANO1, in regulating fluid secretion in glands that produce tears and saliva. These gate-like proteins control how certain ions move in or out of cells, which also influences water movement. Once activated by external stimuli, TRPV4 allows calcium ions to enter the cell, causing ANO1 to open and chloride ions to leave. This results in water also exiting the cell through dedicated channels, before being collected in ducts connected to the outside of the body. TRPV4, which is activated by heat, is also present in human sweat gland cells. This prompted Kashio et al. to examine the role of these channels in sweat production, focusing on mice as well as AIGA patients. Probing TRPV4, ANO1 and AQP5 (a type of water channel) levels using fluorescent antibodies confirmed that these channels are all found in the same sweat gland cells in the foot pads of mice. Further experiments highlighted that TRPV4 mediates sweat production in these animals via ANO1 activation. As rodents do not regulate their body temperature by sweating, Kashio et al. explored the biological benefits of having sweaty paws. Mice lacking TRPV4 had reduced sweating and were less able to climb a slippery slope, suggesting that a layer of sweat helps improve traction. Finally, Kashio et al. compared samples obtained from healthy volunteers with those from AIGA patients and found that TRPV4 levels are lower in individuals affected by the disease. Overall, these findings reveal new insights into the underlying mechanisms of sweating, with TRPV4 a potential therapeutic target for conditions like AIGA. The results also suggest that sweating could be controlled by local changes in temperature detected by heat-sensing channels such as TRPV4. This would depart from our current understanding that sweating is solely controlled by the autonomic nervous system, which regulates involuntary bodily functions such as saliva and tear production.


Subject(s)
Sweating , TRPV Cation Channels , Temperature , Animals , TRPV Cation Channels/metabolism , TRPV Cation Channels/genetics , Mice , Sweating/physiology , Mice, Knockout , Anoctamin-1/metabolism , Anoctamin-1/genetics , Sweat Glands/metabolism , Humans , Male
18.
Mol Ther Oncol ; 32(2): 200818, 2024 Jun 20.
Article in English | MEDLINE | ID: mdl-38966038

ABSTRACT

Bladder cancer (BlCa) is an extensively heterogeneous disease that leads to great variability in tumor evolution scenarios and lifelong patient surveillance, emphasizing the need for modern, minimally invasive precision medicine. Here, we explored the clinical significance of copy number alterations (CNAs) in BlCa. CNA profiling was performed in 15 patient-derived xenografts (PDXs) and validated in The Cancer Genome Atlas BlCa (TCGA-BLCA; n = 408) and Lindgren et al. (n = 143) cohorts. CDKN2A copy number loss was identified as the most frequent CNA in bladder tumors, associated with reduced CDKN2A expression, tumors of a papillary phenotype, and prolonged PDX survival. The study's screening cohort consisted of 243 BlCa patients, and CDKN2A copy number was assessed in genomic DNA and cell-free DNA (cfDNA) from 217 tumors and 189 pre-treatment serum samples, respectively. CDKN2A copy number loss was correlated with superior disease-free and progression-free survival of non-muscle-invasive BlCa (NMIBC) patients. Moreover, a higher CDKN2A index (CDKN2A/LEP ratio) in pre-treatment cfDNA was associated with advanced tumor stage and grade and short-term NMIBC progression to invasive disease, while multivariate models fitted for CDKN2A index in pre-treatment cfDNA offered superior risk stratification of T1/high-grade and EORTC high-risk patients, enhancing prediction of treatment outcome. CDKN2A copy number status could serve as a minimally invasive tool to improve risk stratification and support personalized prognosis in BlCa.

19.
Biochem Biophys Res Commun ; 728: 150324, 2024 Jun 28.
Article in English | MEDLINE | ID: mdl-38968772

ABSTRACT

Ras homolog gene family member C (RhoC) is a GTPase involved in cell migration, implicated in epithelial-mesenchymal transition and treatment resistance and metastasis of cancer. For example, RhoC has been shown to be involved in resistance to radiation in cervical carcinoma. Here, the effect of X-ray irradiation on RhoC expression in prostate cancer (PCa) xenografts was investigated in both xenografts in regression and relapse. Male BALB/cAnNRj-Foxn1nu/nu mice were inoculated with 4-6 million LNCaP-FGC cells and established xenografts were irradiated with X-rays (200 kV, 1 Gymin-1), 5, 10 or 15 Gy using a Gulmay Medical X-ray system. Expression of RhoC and Ki67, a known proliferation marker, was investigated in xenografts, given 15 Gy, 7 days (midst response as measured by size) or 3 weeks (relapse) post irradiation. Staining was quantified using the Halo software (v2.3.2089.34) with the Indica Labs - cytonuclear v1.6 algorithm. RhoC and Ki67 staining was divided into weak, medium, and strong staining and the percentage of cells stained, single and dual staining, was quantified. The HALO software was further used to classify the tissue in each section so that analysis of RhoC and Ki67 expression in cancer cells, stroma and necrotic areas could be done separately. The results showed that RhoC expression in cancer and stroma cells was significantly higher in relapsed xenografts than in those in regression. This was not seen for Ki67 staining, where the percentage of stained cells were the same in regressing and relapsing tumors. RhoC could be a useful biomarker to confirm relapse following external beam radiation therapy.

20.
Biomed Pharmacother ; 177: 117090, 2024 Jul 04.
Article in English | MEDLINE | ID: mdl-38968796

ABSTRACT

Alzheimer's disease (AD) is a progressive neurodegenerative disease accompanied by irreversible cognitive impairment. A deleterious feedback loop between oxidative stress and neuroinflammation in early AD exacerbates AD-related pathology. Platycodon grandiflorum root extract (PGE) has antioxidant and anti-inflammatory effects in several organs. However, the mechanisms underlying the effects of PGE in the brain remain unclear, particularly regarding its impact on oxidative/inflammatory damage and Aß deposition. Thus, we aim to identify the mechanism through which PGE inhibits Aß deposition and oxidative stress in the brain by conducting biochemical and histological analyses. First, to explore the antioxidant mechanism of PGE in the brain, we induced oxidative stress in mice injected with scopolamine and investigated the effect of PGE on cognitive decline and oxidative damage. We also assessed the effect of PGE on reactive oxygen species (ROS) generation and the expressions of antioxidant enzymes and neurotrophic factor in H2O2- and Aß-treated HT22 hippocampal cells. Next, we investigated whether PGE, which showed antioxidant effects, could reduce Aß deposition by mitigating neuroinflammation, especially microglial phagocytosis. We directly verified the effect of PGE on microglial phagocytosis, microglial activation markers, and pro-inflammatory cytokines in Aß-treated BV2 microglial cells. Moreover, we examined the effect of PGE on neuroinflammation, inducing microglial responses in Aß-overexpressing 5XFAD transgenic mice. PGE exerts antioxidant effects in the brain, enhances microglial phagocytosis of Aß, and inhibits neuroinflammation and Aß deposition, ultimately preventing neuronal cell death in AD. Taken together, our findings indicate that the therapeutic potential of PGE in AD is mediated by its targeting of multiple pathological processes.

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