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Current Medicinal Chemistry Jun 2024Oral squamous cell carcinoma (OSCC) represents the primary form of oral cancer, posing a significant global health threat. The existing chemotherapy options are...
BACKGROUND
Oral squamous cell carcinoma (OSCC) represents the primary form of oral cancer, posing a significant global health threat. The existing chemotherapy options are accompanied by notable side effects impacting patient treatment adherence. Consequently, the exploration and development of novel substances with enhanced anticancer effects and fewer side effects have become pivotal in the realms of biological and chemical science.
OBJECTIVE
This work presents the pioneering examples of naphthoquinone-coumarin hybrids as a new category of highly effective cytotoxic substances targeting oral squamous cell carcinoma (OSCC).
METHODS
Given the significance of both naphthoquinones and coumarins as essential pharmacophores/ privileged structures in the quest for anticancer compounds, this study focused on the synthesis and evaluation of novel naphthoquinones/coumarin hybrids against oral squamous cell carcinoma.
RESULTS
By several in vitro, in silico, and in vivo approaches, we demonstrated that compound 6e was highly cytotoxic against OSCC cells and several other cancer cell types and was more selective than current chemotherapeutic drugs (carboplatin) and the naphthoquinone lapachol. Furthermore, compound 6e was non-hemolytic and tolerated in vivo at 50 mg/kg with an LD50 of 62.5 mg/kg. Furthermore, compound 6e did not induce apoptosis and cell cycle arrest but led to intracellular vesicle formation with LC3 aggregation in autophagosomes, suggesting an autophagic cell death. Additionally, 6e had a high-affinity potential for PKM2 protein, higher than the known ligands, such as lapachol or shikonin, and was able to inhibit this enzyme activity in vitro.
CONCLUSION
We assert that compound 6e shows promise as a potential lead for a novel chemotherapeutic drug targeting OSCC, with potential applicability to other cancer types.
PubMed: 38877863
DOI: 10.2174/0109298673298471240605072658 -
Alzheimer's Research & Therapy Jun 2024Our study aims to evaluate the genetic and phenotypic spectrum of Frontotemporal dementia (FTD) gene variant carriers in Chinese populations, investigate mutation...
BACKGROUND
Our study aims to evaluate the genetic and phenotypic spectrum of Frontotemporal dementia (FTD) gene variant carriers in Chinese populations, investigate mutation frequencies, and assess the functional properties of TBK1 and OPTN variants.
METHODS
Clinically diagnosed FTD patients underwent genetic analysis through exome sequencing, repeat-primed polymerase chain reaction, and Sanger sequencing. TBK1 and OPTN variants were biologically characterized in vitro using immunofluorescence, immunoprecipitation, and immunoblotting analysis. The frequencies of genes implicated in FTD in China were analyzed through a literature review and meta-analysis.
RESULTS
Of the 261 Chinese FTD patients, 61 (23.4%) carried potential causative variants in FTD-related genes, including MAPT (n = 17), TBK1 (n = 7), OPTN (n = 6), GRN (n = 6), ANXA11 (n = 4), CHMP2B (n = 3), C9orf72 GGGGCC repeats (n = 2), CYLD (n = 2), PRNP (n = 2), SQSTM1 (n = 2), TARDBP (n = 2), VCP (n = 1), CCNF (n = 1), CHCHD10 (n = 1), SIGMAR1 (n = 1), CHCHD2 (n = 1), FUS (n = 1), TMEM106B (n = 1), and UBQLN2 (n = 1). 29 variants can be considered novel, including the MAPT p.D54N, p.E342K, p.R221P, p.T263I, TBK1 p.E696G, p.I37T, p.E232Q, p.S398F, p.T78A, p.Q150P, p.W259fs, OPTN p.R144G, p.F475V, GRN p.V473fs, p.C307fs, p.R101fs, CHMP2B p.K6N, p.R186Q, ANXA11 p.Q155*, CYLD p.T157I, SQSTM1 p.S403A, UBQLN2 p.P509H, CCNF p.S160N, CHCHD10 p.A8T, SIGMAR1 p.S117L, CHCHD2 p.P53fs, FUS p.S235G & p.S236G, and TMEM106B p.L144V variants. Patients with TBK1 and OPTN variants presented with heterogeneous clinical phenotypes. Functional analysis demonstrated that TBK1 I37T and E232Q mutants showed decreased autophosphorylation, and the OPTN phosphorylation was reduced by the TBK1 I37T mutant. The OPTN-TBK1 complex formation was enhanced by the TBK1 E696G mutant, while OPTN R144G and F475V mutants exhibited reduced recruitment to autophagosomes compared to the wild-type. The overall frequency of TBK1 and OPTN in Chinese FTD patients was 2.0% and 0.3%, respectively.
CONCLUSIONS
Our study demonstrates the extensive genetic and phenotypic heterogeneity of Chinese FTD patients. TBK1 mutations are the second most frequent cause of clinical FTD after MAPT in the Chinese.
Topics: Aged; Female; Humans; Male; Middle Aged; Cell Cycle Proteins; China; East Asian People; Frontotemporal Dementia; Genetic Predisposition to Disease; Membrane Transport Proteins; Mutation; Protein Serine-Threonine Kinases; Transcription Factor TFIIIA
PubMed: 38872230
DOI: 10.1186/s13195-024-01493-w -
The Journal of Clinical Investigation Jun 2024The identification of genes that confer either extension of lifespan or accelerate age-related decline was a step forward in understanding the mechanisms of ageing and...
The identification of genes that confer either extension of lifespan or accelerate age-related decline was a step forward in understanding the mechanisms of ageing and revealed that it is partially controlled by genetics and transcriptional programs. Here we discovered that the human DNA sequence C16ORF70 encoded for a protein, named MYTHO (Macroautophagy and YouTH Optimizer), which controls life- and health-span. MYTHO protein is conserved from C. elegans to humans and its mRNA was upregulated in aged mice and elderly people. Deletion of the ortholog myt-1 gene in C. elegans dramatically shortened lifespan and decreased animal survival upon exposure to oxidative stress. Mechanistically, MYTHO is required for autophagy likely because it acts as a scaffold that binds WIPI2 and BCAS3 to recruit and assemble the conjugation system at the phagophore, the nascent autophagosome. We conclude that MYTHO is a transcriptionally regulated initiator of autophagy that is central in promoting stress resistance and healthy ageing.
PubMed: 38869949
DOI: 10.1172/JCI165814 -
Zhongguo Zhen Jiu = Chinese Acupuncture... Jun 2024To observe the effects of electroacupuncture (EA) on the autophagy of ovarian granulosa cells in rats with premature ovarian insufficiency (POI), and explore the...
OBJECTIVE
To observe the effects of electroacupuncture (EA) on the autophagy of ovarian granulosa cells in rats with premature ovarian insufficiency (POI), and explore the mechanism of EA in improving POI.
METHODS
Thirty-two female SD rats were randomly divided into a blank group (=8) and a model making group (=24). The rats in the model making group were injected intraperitoneally with cyclophosphamide for 15 days to establish the POI model (the dosage on the 1st day was 50 mg/kg, and 8 mg/kg from the 2nd day to 15th day). The successfully modeled rats were then randomly divided into a model group, an EA group, and an estradiol (E2) group, with 8 rats in each group. Rats in the EA group received EA at bilateral "Gongsun" (SP 4) with continuous wave, frequency of 2 Hz, and current intensity of 0.1 to 1 mA, 20 min per treatment, once daily for 14 days. Rats in the E2 group were administered with E2 (0.01 mg/mL) by gavage (10 mL/kg), once daily for 14 days. The changes in estrous cycle were observed by rapid Giemsa staining before and after modeling. After intervention, ovarian tissue morphology was observed by HE staining; serum levels of follicle-stimulating hormone (FSH), E2, anti-Mullerian hormone (AMH), and inhibin B (INHB) were detected by ELISA; immunofluorescence staining was used to observe the expression of p62 in ovarian granulosa cells; the ultrastructure of ovarian granulosa cells was observed by transmission electron microscopy, and the number of autophagosomes and autolysosomes was compared; Western blot and real-time fluorescence quantitative PCR were used to detect the protein and mRNA expression of p62, Beclin-1, and microtubule-associated protein 1A/1B-light chain 3 (LC3) in ovarian tissue.
RESULTS
The results of vaginal smears in the blank group showed regular cyclical changes; the rats in the model group showed prolonged estrous cycle or cycle arrest, mostly in proestrus or metestrus, with overall ovarian atrophy, disordered structure, and decreased granulosa cells. Compared with the blank group, rats in the model group showed increased serum FSH level (<0.01), decreased serum levels of E2, AMH, and INHB (<0.01), decreased positive expression of p62 in ovarian granulosa cells (<0.01), with obvious swelling of ovarian granulosa cells, mild to moderate swelling of mitochondria, slight expansion of rough endoplasmic reticulum, and hypertrophy of Golgi apparatus; the number of autophagosomes and autolysosomes in the ovaries was increased (<0.01), the expression of p62 protein and mRNA was decreased (<0.01), and the expression of Beclin-1 and LC3 protein and mRNA in ovarian tissue was increased (<0.01). Compared with the model group, rats in the EA group and the E2 group showed decreased serum FSH levels (<0.01), increased levels of E2, AMH, and INHB (<0.01), increased positive expression of p62 in ovarian granulosa cells (<0.01), alleviated degree of ovarian granulosa cell damage, with relatively intact organelle morphology, and decreased number of autophagosomes and autolysosomes in the ovaries (<0.01); the rats also showed increased expression of p62 protein and mRNA (<0.01), and decreased expression of Beclin-1 and LC3 protein and mRNA (<0.01) in ovarian tissue.
CONCLUSION
EA at "Gongsun" (SP 4) could improve ovarian reserve function in POI rats by reducing the number of autophagosomes and autolysosomes, up-regulating p62 expression, and down-regulating Beclin-1 and LC3 expression, thus inhibiting autophagy of ovarian granulosa cells, and regulating the serum levels of FSH, E2, AMH, and INHB.
Topics: Animals; Female; Electroacupuncture; Primary Ovarian Insufficiency; Rats; Autophagy; Rats, Sprague-Dawley; Humans; Granulosa Cells; Disease Models, Animal
PubMed: 38867630
DOI: 10.13703/j.0255-2930.20230930-k0001 -
Frontiers in Immunology 2024Dengue virus (DENV), transmitted by infected mosquitoes, is a major public health concern, with approximately half the world's population at risk for infection. Recent...
Dengue virus (DENV), transmitted by infected mosquitoes, is a major public health concern, with approximately half the world's population at risk for infection. Recent decades have increasing incidence of dengue-associated disease alongside growing frequency of outbreaks. Although promising progress has been made in anti-DENV immunizations, post-infection treatment remains limited to non-specific supportive treatments. Development of antiviral therapeutics is thus required to limit DENV dissemination in humans and to help control the severity of outbreaks. Dendritic cells (DCs) are amongst the first cells to encounter DENV upon injection into the human skin mucosa, and thereafter promote systemic viral dissemination to additional human target cells. Autophagy is a vesicle trafficking pathway involving the formation of cytosolic autophagosomes, and recent reports have highlighted the extensive manipulation of autophagy by flaviviruses, including DENV, for viral replication. However, the temporal profiling and function of autophagy activity in DENV infection and transmission by human primary DCs remains poorly understood. Herein, we demonstrate that mechanisms of autophagosome formation and extracellular vesicle (EV) release have a pro-viral role in DC-mediated DENV transmission. We show that DENV exploits early-stage canonical autophagy to establish infection in primary human DCs. DENV replication enhanced autophagosome formation in primary human DCs, and intrinsically-heightened autophagosome biogenesis correlated with relatively higher rates of DC susceptibility to DENV. Furthermore, our data suggest that viral replication intermediates co-localize with autophagosomes, while productive DENV infection introduces a block at the late degradative stages of autophagy in infected DCs but not in uninfected bystander cells. Notably, we identify for the first time that approximately one-fourth of DC-derived CD9/CD81/CD63+ EVs co-express canonical autophagy marker LC3, and demonstrate that DC-derived EV populations are an alternative, cell-free mechanism by which DCs promote DENV transmission to additional target sites. Taken together, our study highlights intersections between autophagy and secretory pathways during viral infection, and puts forward autophagosome accumulation and viral RNA-laden EVs as host determinants of DC-mediated DENV infection in humans. Host-directed therapeutics targeting autophagy and exocytosis pathways thus have potential to enhance DC-driven resistance to DENV acquisition and thereby limit viral dissemination by initial human target cells following mosquito-to-human transmission of DENV.
Topics: Humans; Dengue Virus; Dendritic Cells; Autophagy; Dengue; Autophagosomes; Secretory Pathway; Virus Replication; Extracellular Vesicles; Cells, Cultured
PubMed: 38863700
DOI: 10.3389/fimmu.2024.1260439 -
Frontiers in Cellular Neuroscience 2024Transforming growth factor β1 (TGF-β1) has a neuroprotective function in traumatic brain injury (TBI) through its anti-inflammatory and immunomodulatory properties....
Transforming growth factor β1 (TGF-β1) has a neuroprotective function in traumatic brain injury (TBI) through its anti-inflammatory and immunomodulatory properties. However, the precise mechanisms underlying the neuroprotective actions of TGF-β1 on the cortex require further investigation. In this study, we were aimed to investigate the regulatory function of TGF-β1 on neuronal autophagy and apoptosis using an primary cortical neuron trauma-injury model. LDH activity was assayed to measure cell viability, and intracellular [Ca] was measured using Fluo-4-AM in an primary cortical neuron trauma-injury model. RNA-sequencing (RNAseq), immunofluorescent staining, transmission electron microscopy (TEM), western blot and CTSD activity detection were employed. We observed significant enrichment of DEGs related to autophagy, apoptosis, and the lysosome pathway in trauma-injured cortical neurons. TEM confirmed the presence of autophagosomes as well as autophagolysosomes. Western blot revealed upregulation of autophagy-related protein light chain 3 (LC3-II/LC3-I), sequestosome 1 (SQSTM1/p62), along with apoptosis-related protein cleaved-caspase 3 in trauma-injured primary cortical neurons. Furthermore, trauma-injured cortical neurons showed an upregulation of lysosomal marker protein (LAMP1) and lysosomal enzyme mature cathepsin D (mCTSD), but a decrease in the activity of CTSD enzyme. These results indicated that apoptosis was up-regulated in trauma- injured cortical neurons at 24 h, accompanied by lysosomal dysfunction and impaired autophagic flux. Notably, TGF-β1 significantly reversed these changes. Our results suggested that TGF-β1 exerted neuroprotective effects on trauma- injured cortical neurons by reducing lysosomal dysfunction, decreasing the accumulation of autophagosomes and autophagolysosomes, and enhancing autophagic flux.
PubMed: 38863498
DOI: 10.3389/fncel.2024.1381279 -
Nature Communications Jun 2024Cancer-associated fibroblasts (CAFs) have emerged as a dominant non-hematopoietic cell population in the tumour microenvironment, serving diverse functions in tumour...
Cancer-associated fibroblasts (CAFs) have emerged as a dominant non-hematopoietic cell population in the tumour microenvironment, serving diverse functions in tumour progression. However, the mechanisms via which CAFs influence the anti-tumour immunity remain poorly understood. Here, using multiple tumour models and biopsies from cancer patients, we report that α-SMA CAFs can form immunological synapses with Foxp3 regulatory T cells (Tregs) in tumours. Notably, α-SMA CAFs can phagocytose and process tumour antigens and exhibit a tolerogenic phenotype which instructs movement arrest, activation and proliferation in Tregs in an antigen-specific manner. Moreover, α-SMA CAFs display double-membrane structures resembling autophagosomes in their cytoplasm. Single-cell transcriptomic data showed an enrichment in autophagy and antigen processing/presentation pathways in α-SMA-expressing CAF clusters. Conditional knockout of Atg5 in α-SMA CAFs promoted inflammatory re-programming in CAFs, reduced Treg cell infiltration and attenuated tumour development. Overall, our findings reveal an immunosuppressive mechanism entailing the formation of synapses between α-SMA CAFs and Tregs in an autophagy-dependent manner.
Topics: T-Lymphocytes, Regulatory; Cancer-Associated Fibroblasts; Humans; Immunological Synapses; Animals; Tumor Microenvironment; Mice; Autophagy; Actins; Autophagy-Related Protein 5; Neoplasms; Mice, Inbred C57BL; Forkhead Transcription Factors; Female; Mice, Knockout
PubMed: 38862534
DOI: 10.1038/s41467-024-49282-1 -
Scientific Reports Jun 2024Lung cancer is the most common oncological disease worldwide, with non-small cell lung cancer accounting for approximately 85% of lung cancer cases. α-Hederin is a...
Lung cancer is the most common oncological disease worldwide, with non-small cell lung cancer accounting for approximately 85% of lung cancer cases. α-Hederin is a monodesmosidic triterpenoid saponin isolated from the leaves of Hedera helix L. or Nigella sativa and has been extensively studied for its antitumor activity against a variety of tumor cells. It has been suggested that α-Hederin is a potential regulator of autophagy and has high promise for application. However, the specific mechanism and characteristics of α-Hederin in regulating autophagy are not well understood. In this study, we confirmed the potential of α-Hederin application in lung cancer treatment and comprehensively explored the mechanism and characteristics of α-Hederin in regulating autophagy in lung cancer cells. Our results suggest that α-Hederin is an incomplete autophagy inducer that targets mTOR to activate the classical autophagic pathway, inhibits lysosomal acidification without significantly affecting the processes of autophagosome transport, lysosome biogenesis, autophagosome and lysosome fusion, and finally leads to impaired autophagic flux and triggers autophagic damage in NSCLC.
Topics: Humans; Carcinoma, Non-Small-Cell Lung; Lysosomes; Autophagy; Lung Neoplasms; Oleanolic Acid; Saponins; Cell Line, Tumor; TOR Serine-Threonine Kinases; Autophagosomes; A549 Cells
PubMed: 38858422
DOI: 10.1038/s41598-024-63348-6 -
Autophagy Jun 2024The serine/threonine kinase, PINK1, and the E3 ubiquitin ligase, PRKN/Parkin facilitate LC3-dependent autophagosomal encasement and lysosomal clearance of dysfunctional...
The serine/threonine kinase, PINK1, and the E3 ubiquitin ligase, PRKN/Parkin facilitate LC3-dependent autophagosomal encasement and lysosomal clearance of dysfunctional mitochondria, and defects in this pathway contribute to the pathogenesis of numerous cardiometabolic and neurological diseases. Although dynamic actin remodeling has recently been shown to play an important role in governing spatiotemporal control of mitophagy, the mechanisms remain unclear. We recently found that the RhoGAP, ARHGAP26/GRAF1 is a PRKN-binding protein that is rapidly recruited to damaged mitochondria where upon phosphorylation by PINK1 it serves to coordinate phagophore capture by regulating mitochondrial-associated actin remodeling and by facilitating PRKN-LC3 interactions. Because ARHGAP26 phosphorylation on PINK1-dependent sites is dysregulated in human heart failure and ARHGAP26 depletion in mouse hearts blunts mitochondrial clearance and attenuates compensatory metabolic adaptations to stress, this enzyme may be a tractable target to treat the many diseases associated with mitochondrial dysfunction.
PubMed: 38855880
DOI: 10.1080/15548627.2024.2361576 -
Experimental Cell Research Jul 2024Autophagy phenomenon in the cell maintains proteostasis balance by eliminating damaged organelles and protein aggregates. Imbalance in autophagic flux may cause...
Autophagy phenomenon in the cell maintains proteostasis balance by eliminating damaged organelles and protein aggregates. Imbalance in autophagic flux may cause accumulation of protein aggregates in various neurodegenerative disorders. Regulation of autophagy by either calcium or chaperone play a key role in the removal of protein aggregates from the cell. The neuromuscular rare genetic disorder, GNE Myopathy, is characterized by accumulation of rimmed vacuoles having protein aggregates of β-amyloid and tau that may result from altered autophagic flux. In the present study, the autophagic flux was deciphered in HEK cell-based model for GNE Myopathy harbouring GNE mutations of Indian origin. The refolding activity of HSP70 chaperone was found to be reduced in GNE mutant cells compared to wild type controls. The autophagic markers LC3II/I ratio was altered with increased number of autophagosome formation in GNE mutant cells compared to wild type cells. The cytosolic calcium levels were also increased in GNE mutant cells of Indian origin. Interestingly, treatment of GNE mutant cells with HSP70 activator, BGP-15, restored the expression and refolding activity of HSP70 along with autophagosome formation. Treatment with calcium chelator, BAPTA-AM restored the cytoplasmic calcium levels and autophagosome formation but not LC3II/I ratio significantly. Our study provides insights towards GNE mutation specific response for autophagy regulation and opens up a therapeutic advancement area in calcium signalling and HSP70 function for GNE related Myopathy.
Topics: Humans; Autophagy; Mutation; Calcium; Distal Myopathies; HSP70 Heat-Shock Proteins; Multienzyme Complexes; HEK293 Cells; Autophagosomes; India
PubMed: 38852763
DOI: 10.1016/j.yexcr.2024.114118