Restoring mitochondrial homeostasis to inhibit apoptosis in renal tubular epithelial cells (RTECs) has emerged as a promising therapeutic strategy for diabetic kidney disease (DKD). This study focuses on the therapeutic effect and mechanism of the triterpenoid compound cycloastragenol (CAG) from Astragali Radix in the treatment of DKD. The DKD model was established in C57BL/6Jdb/db mice and AGEs-induced HK-2 cells. Various biological techniques such as WB and RT-PCR revealed that CAG enhanced mitophagy via TFEB, reducing apoptosis in RTECs. Mechanistic studies combining CETSA, molecular docking, and molecular dynamics simulations confirmed the CAG-ERK interaction. CAG improved renal function and reduced renal tubular injury in db/db mice. CAG effectively reduced the accumulation of mitoROS, enhanced mitochondrial membrane potential, promoted mitophagy and mitochondrial biogenesis, and restored mitochondrial homeostasis. Mechanistically, CAG enhanced mitophagy in db/db mice and AGEs-induced HK-2 cells by stimulating the autophagic flux via regulating TFEB. Moreover, CAG inhibited AGEs-induced HK-2 apoptosis, which was reversed by autophagy inhibitor chloroquine (CQ) and siRNA-TFEB. Importantly, after mutating the valine (VAL) at position 39 of the ERK to alanine (ALA), the binding effect between CAG and ERK was significantly reduced, revealing that CAG directly bound ERK at 39VAL, inhibiting its phosphorylation, thus preventing the phosphorylation of the S142 site of TFEB and enabling TFEB to translocate into the nucleus. CAG ameliorated renal tubule damage in DKD by regulating mitochondrial quality though targeting ERK to regulate TFEB. This research advances drug development and proposes lifestyle interventions (e.g., dietary supplements).

The role and mechanism of autophagy-regulated Keap1/Nrf2 pathway in oxidative stress-induced cartilage endplate cell apoptosis.

TRIM21 knockdown suppresses ESCC progression and induces autophagy-mediated apoptosis via AKT/mTOR signaling pathway.

Blocking T-type calcium channels disrupts spermatogenesis in vivo and adversely affects spermatocytes in vitro by impairing mitochondrial function and autophagic flux.

A graphene oxide assisted surface plasmon resonance sensor for chloroquine phosphate detection: A theoretical and experimental study

Bi3O4Br/Ce-based-MOF Z-scheme heterojunction enhanced chloroquine phosphate degradation under visible light irradiation: Key role of targeted adsorption-photocatalytic synergistic effect

Stage-Dependent Expression and Vacuolar Localization of lasmodium berghei Chloroquine Resistance Transporter (CRT).

Computational investigation of mutations in PfCRT and PfDHFR proteins for emerging resistance of Plasmodium falciparum to antimalarial drugs.

Methionine addiction is a metabolic hallmark of cancer. Recombinant methioninase (rMETase) targets methionine addiction and effectively depletes methionine. rMETase has shown synergy with chemotherapeutic agents on numerous types of cancer cells. Chloroquine (CQ), an anti-autophagy agent, has demonstrated anti-cancer efficacy in pre-clinical studies. The present study aimed to evaluate the cancer selectivity and synergistic efficacy of rMETase and CQ in a co-culture model of colon-cancer cells and normal fibroblasts. HCT116 human colon-cancer cells and Hs-27 human normal fibroblasts were co-cultured in Dulbecco's Modified Eagle's Medium (DMEM) and treated with rMETase (0.1-0.5 U/ml) alone, CQ (10-60 μM) alone, or rMETase at various concentrations in combination with CQ (20 μM). Cell morphology and viability were monitored for six days using phase-contrast microscopy (Olympus IX71). The effects of each treatment on the cancer cells and normal fibroblasts were compared. rMETase treatment selectively reduced HCT116 viability in a dose-dependent manner while sparing normal fibroblasts. In contrast, high-concentrations of CQ decreased the viability of both cell types, with strong cytotoxicity at ≥40 μM. Combination treatment with rMETase and low-dose CQ (20 μM) produced greater selective efficacy against the cancer cells than rMETase alone, eliminating the cancer cells and without significant inhibition of fibroblast viability. rMETase has selective efficacy against cancer cells in the presence of normal cells, and its efficacy is significantly enhanced selectively on the cancer cells by CQ. The results of the present study suggest the potential for future clinical application of the combination of rMETase and CQ for cancer treatment.

Chloroquine (CQ), a well-known antimalarial agent, has been proposed as a potential antiviral compound due to its ability to interfere with multiple cellular pathways critical for viral replication. Although CQ exhibits broad-spectrum antiviral activity, its effect on host innate immune responses remains incompletely understood. The timing of CQ administration, whether before or after infection, may lead to different immunological outcomes. Therefore, the immunomodulatory effects of CQ should be carefully evaluated before antiviral therapy. To investigate the immunomodulatory role of CQ (50 M), we used immunofluorescence staining, western blotting, and reporter assays to evaluate innate immune activation in A549 cells. We established a doxycycline-inducible system to activate MAVS-mediated signaling without viral infection. Plaque assays and antiviral tests were performed to measure viral replication, while cytokine array and RT-qPCR were used to quantify cytokine production. Mitochondrial morphology was assessed using immunofluorescence microscopy. CQ enhanced innate immune responses triggered by dengue virus infection and Poly(I:C) stimulation. This enhancement was associated with the activation of the mitochondrial antiviral signaling protein (MAVS) and its upstream receptors, including retinoic acid-inducible gene I and melanoma differentiation-associated protein 5. CQ strengthened MAVS-dependent antiviral signaling and increased IL-6 induction more than 13-fold. Alterations in mitochondrial morphology may contribute to this immunostimulatory effect. CQ promotes MAVS-mediated antiviral and inflammatory cytokine responses, potentially through its effect on mitochondrial dynamics. These findings indicate that while CQ may enhance antiviral defense, its immune-stimulating properties should be carefully evaluated prior to its use as an antiviral agent in treating RNA virus infections.

To develop a novel multifunctional nanoparticle platform by combining mesenchymal stem cell-derived nanovesicles (MSC-NVs) with poly(lactic-coglycolic acid) (PLGA) nanoparticles for Alzheimer's disease (AD) therapy. Mesenchymal stem cell-derived nanovesicle-poly(lactic-coglycolic acid) nanoparticles (MSC-PLGA-NPs) were prepared via sonication-loading. Blood-brain barrier (BBB) penetration was evaluated using in vitro transwell models and in vivo mouse models. Lysosomal function, autophagy, pathological protein clearance, and anti-inflammatory effects were assessed using various cellular and molecular biology techniques. MSC-PLGA-NPs demonstrated 2.3-fold higher BBB penetration efficiency compared to PLGA alone. In a chloroquine(CQ)-induced lysosomal injury model and mice model, they effectively restored lysosomal pH, enhanced autophagy (reducing LC3-II/I ratio by 0.4-fold and p62 expression by 52%), cleared amyloid precursor protein (APP) and phosphorylated tau (p-tau) proteins, and inhibited IL-6 and TNF-α without hepatorenal toxicity. These results demonstrate that MSC-PLGA-NPs, a novel multifunctional nanoparticle platform, synergistically integrates the BBB penetration capability of MSC-NVs and the lysosomal acidification function of PLGA. The synergistic combination represents a pioneering "delivery-repair-clearance" integrated strategy for AD therapy. Offering significant advantages over single-component approaches, MSC-PLGA-NPs provide a promising preclinical candidate and new insight into lysosome-targeted nanomedicines for neurodegenerative diseases.

Oral cancer, the most common head and neck malignancy, has a high recurrence rate and poor prognosis largely owing to chemotherapy resistance. The adverse effects of conventional therapies have prompted investigations into safer and more effective alternative therapies. Chloroquine (CQ) and hydroxychloroquine (HCQ) have shown potential owing to their roles in autophagy modulation and immune regulation. This study clarifies the selective efficacy of hydroxychloroquine (HCQ) and chloroquine (CQ) in oral squamous cell carcinoma models, emphasizing distinct responses in gingival (Ca9-22) and tongue (SCC-9) carcinoma cells. Non-oncogenic oral epithelial cells (GMSM-K) and oral carcinoma cell lines from the tongue (SCC-9, Cal-27) and gingiva (Ca9-22) were used. Cell viability, cytotoxicity, and colony formation were assessed via MTT, LDH, and crystal violet assays. Flow cytometry was used to measure apoptosis, autophagy, oxidative stress, mitochondrial membrane potential, and DNA damage. The transcriptomic profiles of apoptosis and autophagy-related genes were assessed by qPCR arrays. Bioinformatics analysis allowed estimation of the main gene interaction networks. Pre-screening showed that GMSM-K and Cal-27 cells were non-responsive or exhibited non-specific toxicity at high doses; therefore, subsequent analyses focused on Ca9-22 (GC) and SCC-9 (TC). HCQ significantly reduced viability and colony formation in Ca9-22 cells while moderately affecting SCC-9 cells. Autophagy inhibition was accompanied by compensatory up-regulation of autophagy-related genes, consistent with feedback activation of TFEB and FOXO3a pathways. Gene expression profiling and flow-cytometry analyses revealed cell-type-specific differences in apoptosis, mitochondrial potential, and DNA damage, suggesting HCQ’s selective anti-tumor potential in gingival carcinoma. These findings highlight HCQ as a repurposed adjuvant therapy that modulates autophagy and apoptosis to enhance chemosensitivity in oral cancer.

Malaria is a deadly disease for which therapeutic options are threatened by the rise of antimalarial resistance. Inhibiting the formation of hemozoin (the product of heme detoxification) in the digestive vacuole (DV) is the mechanism of action of numerous antimalarial drugs, including those in development as new therapies. This drug target remains attractive as hemozoin is an abiotic and non-mutable molecule, unique to the parasite. The underlying parasite biology of the heme detoxification pathway is complex and requires a deeper understanding. This study focuses on the DV of Plasmodium falciparum, utilizing confocal microscopy, immunofluorescence, immunoblotting and cellular fractionation techniques to study its native state over time. Using parameters such as the uptake into and growth of the DV, relative abundance of plasmepsins (PMs) I and IV as well as basal levels of hemoglobin, heme and hemozoin, it was found that DV physiology in chloroquine (CQ)-sensitive NF54 follows three distinct developmental phases: the lag-type growth (20 to 28 h), rapid growth phase (28 to 40 h) and the plateau (40 to 48 h). These phases hold specific characteristics with respect to the investigated parameters. In addition, key differences between CQ-sensitive NF54 and CQ-resistant Dd2 parasites were observed.

Malaria infection induces a cascade of secondary pathological events, among which oxidative stress and inflammation are interconnected. This interaction often exacerbates tissue damage and significantly contributes to disease progression. Given the reported antioxidant property of red palm oil, this study evaluated the therapeutic potential of a red palm oil supplemented diet in mitigating oxidative stress in a murine model of plasmodium berghei infection concurrently administered with standard anti-malaria drugs. Mice weighing 19 ± 2.5g were distributed into seven groups of fifteen (15) animals each for this study. The mice were infected with NK65 Plasmodium berghei strain. The individual effects of red palm oil supplemented diet and of anti-malaria drugs (amodiaquine (AMQ), and Chloroquine (CQ) along with their contributory effects were evaluated. The mice were fed with standard rat chow with 15% v/w red palm oil. The trace elements Selenium (Se), Copper (Cu), Zinc (Zn) and Vitamins A, C and E with its fractions were also determined spectrophotometrically. The studies revealed that trace elements such as Se, Zn, Fe and Cu showed no significant difference between the control and the test groups. Vitamin A showed a statistically significant difference in all test groups compared to the control. Palm oil supplemented diet provided more vitamin A in circulation except group F where the anti-malaria chloroquine reduces the availability of vitamin A in circulation. Palm oil supplemented diet also increased the availability of vitamin C in circulation while chloroquine increased the vitamin level higher in control. Vitamin E levels were markedly increased in the test group compared to the control. Total β-and γ-tocopherol levels were increased significantly except for γ-tocopherol where the level remained increased even with chloroquine treatment. Levels of total tocotrienol, β-and γ-tocotrienol increased significantly except for the group receiving chloroquine treatment. The results obtained have not necessarily shown that red palm oil modulated trace elements during oxidative stress but showed increased availability of antioxidant vitamins in circulation.

Investigate the cellular response of human nucleus pulposus (HNP) cells to serum deprivation, focusing on the role of high mobility group box1(HMGB1) in regulating autophagy and apoptosis, and elucidate the time-dependent activation of autophagy shifting toward apoptosis under nutrient stress. Additionally, the study evaluated the impact of autophagy inhibition by chloroquine (CQ) on apoptosis progression. HNP samples were obtained from the human biobank with exemption from IRB screening (IRB number DC25SASI0012) to evaluate the impact of nutritional deprivation. Comprehensive analyses encompassed detailed evaluations of cellular morphology, viability, DNA integrity, and metabolic function, providing an integrated view of cellular status. Western blotting (WB), fluorescence-activated cell sorting (FACS), and immunofluorescence (IF) were used to detect LC3, P62, HMGB1, and cleaved caspase-3. Real-time quantitative polymerase chain reaction (RT-qPCR) further revealed changes in gene expression related to autophagy (LC3, P62) and apoptosis (caspase-3), highlighting cellular stress responses. Serum deprivation markedly reduced HNP cell viability, altered morphology, and suppressed metabolic activity, while inducing a time-dependent increase in autophagy, peaking at 48 h. Furthermore, elevated LC3-II, decreased P62, and increased cytoplasmic translocation of HMGB1 indicate activation of HMGB1-mediated autophagy. Simultaneously, cleaved caspase-3 levels rose, suggesting HMGB1’s involvement in shifting the balance toward apoptosis. IF and RT-qPCR confirmed enhanced LC3 and cleaved caspase-3 expression, while FACS analysis revealed increased apoptotic cell populations with declining serum levels. These findings highlight a crucial interplay between autophagy and apoptosis regulated by HMGB1 under nutrient-deprived conditions. Eventually, CQ treatment inhibited autophagic flux by blocking LC3-II degradation, thereby amplifying apoptosis. Serum deprivation potently induced HMGB1-mediated autophagy-apoptosis interplay in HNP cells, with CQ enhancing apoptosis by inhibiting autophagy. Supplementary InformationThe online version contains supplementary material available at 10.1038/s41598-025-23026-7.

ObjectiveTo investigated the pathogenic mechanism of NFIX frameshift mutations in Malan syndrome.MethodsReviewed the clinical diagnosis and treatment processes of the Malan syndrome proband, analyzing the relationship between NFIX frameshift mutation genotypes and clinical phenotypes, and the inheritance pattern. To analyzed the functional domain where the mutation was located and the conservation of the mutated amino acid residue, thereby elucidating the potential impact of the mutation on the protein. Validated effects on pre-mRNA splicing using RDDCSC, SpliceAI, and FF databases. Assessed variant pathogenicity via MutationTaster, PolyPhen-2, and VarCards. Constructed wild-type/mutant plasmids, transfected to HEK293T cells, and quantified NFIX mRNA and protein expression levels via qPCR and Western blot. Analyzed degradation pathways using ubiquitin-proteasome inhibitor MG132 and autophagy-lysosome inhibitor Chloroquine (CQ).ResultsThe proband exhibited intellectual disability, distinctive facial features, ocular abnormalities, scoliosis, and primary infertility. A de novo mutation in NFIX (c.164delC, p.Ala55Glyfs*2) associated with these phenotypes was identified. Neither the proband’s father nor his mother was found to have this mutation. Parental testing confirmed de novo inheritance. The amino acid at position 55 was highly conserved and had been Alanine in 5 species. Results from databases including RDDCSC, SpliceAI, and FF indicated that the NFIX c.164delC p.Ala55Glyfs*2 mutation did not affect splicing function. Predictions by MutationTaster and PolyPhen-2 classified the c.707G>A p.Arg236Gln mutation as “damaging,” suggesting an altered amino acid sequence, frameshift mutation, NMD, and potential modification of protein characteristics. Quantitative real-time PCR (qPCR) analysis detected comparable mRNA levels between mutant and wild-type strains. In contrast, Western blotting revealed significantly diminished protein expression in the mutant (P < 0.05), suggesting post-transcriptional regulation effects. Results from protein degradation pathway analysis demonstrated that the truncated protein generated after mutation was degraded via the ubiquitin-proteasome pathway.ConclusionThe NFIX c.164delC p.Ala55Glyfs*2 frameshift mutation did not significantly affect mRNA expression levels, but induced protein degradation via the ubiquitin-proteasome pathway, resulting in haploinsufficiency and ultimately causing Malan syndrome.

ObjectiveTo establish a short-term high-fat/high-cholesterol (HFHC) diet-induced Metabolic dysfunction-associated steatotic liver disease (MASLD) mouse model, and evaluate the effects of rapamycin (RaPa) and chloroquine (CQ) on this model to explore their therapeutic potential and side effects.MethodsAn early MASLD mouse model was constructed via short-term HFHC diet feeding. Model mice were intraperitoneally injected with RaPa or CQ. Drug effects were analyzed on body weight, liver weight, lipid metabolism-related genes (APOB, FASN, PLIN2), inflammatory factors (IL-6, IL-10), and fibrosis markers (LOX, Col-1α-1, CCL2, TGFβ1, PDGFRβ, α-SMA) at mRNA and protein levels.ResultsRaPa ameliorated body weight and liver weight in early MASLD mice, downregulated FASN and PLIN2 expression, upregulated IL-10 mRNA levels, and alleviated hepatic steatosis, but induced metabolic disorders such as Insulin resistance and hyperlipidemia. In contrast, CQ promoted FASN and PLIN2 expression, exacerbated hepatic steatosis, reduced IL-10 mRNA levels, and upregulated fibrosis-related markers (LOX, TGFβ1, PDGFRβ, α-SMA) at both mRNA and protein levels, thereby driving MASLD progression to liver fibrosis. Notably, CQ improved metabolic abnormalities in model mice, including obesity, hyperlipidemia, and Insulin resistance.ConclusionRaPa and CQ exhibit dual effects on early MASLD: RaPa alleviates hepatic steatosis but exacerbates metabolic disorders, whereas CQ improves metabolic abnormalities but accelerates liver fibrosis. This paradox highlights the need to balance metabolic regulation and liver injury prevention in MASLD treatment, providing critical experimental insights for targeted drug development.

Characterization of genes involved in hemoglobin degradation in Plasmodium vivax isolates from Chennai, India, and species of non-human primate malaria.

Ultraviolet/peracetic acid for degradation of antiviral drugs and control of disinfection risk.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) acts as a double-edged sword in cancer by enhancing both anti- and pro-tumorigenic immune cells. In this study, two oncolytic adenoviruses were engineered to modulate GM-CSF expression using different strategies: one with the CMV promoter (oAd-CMV-GM-CSF) and the other using the endogenous viral E3 promoter (oAd-GM-CSF). The impacts of these modifications on transgene expression, cytotoxicity, viral replication, and apoptosis were assessed both in vitro and in vivo. The results demonstrated that oAd-CMV-GM-CSF produced significantly lower GM-CSF levels than oAd-GM-CSF, interestingly oAd-CMV-GM-CSF exhibited increased cytotoxicity and apoptosis compared to oAd-GM-CSF and control groups. The further study showed oAd-CMV-GM-CSF induced profound autophagy through the activation of the Janus kinase 2/Signal Transducer and Activator of Transcription 2 (JAK2/STAT2) signaling pathway. The use of autophagy and JAK-2 inhibitors, Chloroquine (CQ) and AG-490, respectively, significantly mitigated the apoptosis induced by oAd-CMV-GM-CSF. In addition, oAd-CMV-GM-CSF presented a faster viral replication and production of more active progeny virus than oAd-GM-CSF, which could be inhibited by CQ. oAd-CMV-GM-CSF augments propagation of the progeny viruses and induces immunogenic cell death(ICD) in A549 and PANC-1 cells. In vivo oAd-CMV-GM-CSF had stronger anti-tumor effect than oAd-GM-CSF in immunodeficient model and immune-competent model. Our findings indicate that oAd-CMV-GM-CSF induces more profound autophagy and promoting viral replication to enhance the anti-tumor efficacy.