Lysosomal Degradation Pathway Research Articles

Biochemical studies and expression of Atg7/LC3 autophagy genes in mephalan-induced testicular dysfuntion

Autophagy is an intracellular lysosomal degradation pathway and plays a very important role in maintaining intracellular homeostasis. This present study demonstrates the biochemical status and expression of ATG7/LC3 autophagic genes in Melphalan-induced testicular dysfunction. Twenty make Swiss albino mice were maintained under standard conditions of humidity (50 ± 5%), temperature (25 ± 2° C) with free access to food and water. The mice were divided into five groups (n=5 each) and treated by intraperitoneal injection as follows; Group I: vehicle-treated control; Group II: 1 mg/kg/bwt MEP; Group III: 3mg/kg/bwt MEP; Group IV: 5mg/kg/bwt MEP. Histopathological and histochemical evaluation by light microscopy, biochemical assays and sperm parameters evaluation are the various investigation depicted in this study. Result shows a rise percentage of DNA damage and teratozoospermia Index (TZI); decrease of protein concentration and total antioxidant capacity (TAC). Testosterone concentration was depleted. The result also shows a significant decrease of Luteinizing Hormone (LH) level across the groups when compared to the control. This study confirms oxidative stress by increased ROS and MDA levels in the testes of the melphalan-treated mice and defective antioxidants response as evident from diminished activities of antioxidant enzymes. The autophagic gene markers showed an upstream Atg7 gene and downstream of LC3-II/LC3-I ratio. Autophagy is mediated by the expression of Atg7 and LC3 genes and these varying mRNA expression led to autophagic defect.

Frontispiz: Aptamer‐LYTACs for Targeted Degradation of Extracellular and Membrane Proteins

Aptamers. In their Communication (e202218106), Zhi Zhu et al. report the development of aptamer-LYTACs (lysosome targeting chimeras) for the efficient degradation of the platelet-derived growth factor and the membrane protein PTK7 through the lysosomal degradation pathway.

Frontispiece: Aptamer‐LYTACs for Targeted Degradation of Extracellular and Membrane Proteins

Aptamers. In their Communication (e202218106), Zhi Zhu et al. report the development of aptamer-LYTACs (lysosome targeting chimeras) for the efficient degradation of the platelet-derived growth factor and the membrane protein PTK7 through the lysosomal degradation pathway.

Zwitterionic Pluronic analog-coated PLGA nanoparticles for oral insulin delivery

In recent years, zwitterionic materials have drawn great attention in oral drug delivery system due to their capacity for rapid mucus diffusion and enhanced cellular internalization. However, zwitterionic materials tend to show strong polarity that was hard to directly coat hydrophobic nanoparticles (NPs). Inspired by Pluronic coating, a simple and convenient strategy to coat NPs with zwitterionic materials using zwitterionic Pluronic analogs was developed in this investigation. Poly(carboxybetaine)-poly(propylene oxide)-Poly(carboxybetaine) (PCB-PPO-PCB, PPP), containing PPO segments with MW > 2.0 kDa, can effectively adsorb on the surface of PLGA NPs with typical core-shell spherical in shape. The PLGA@PPP4K NPs were stable in gastrointestinal physiological environment and sequentially conquered mucus and epithelium barriers. Proton-assisted amine acid transporter 1 (PAT1) was verified to contribute to the enhanced internalization of PLGA@PPP4K NPs, and the NPs could partially evade lysosomal degradation pathway and utilize retrograde pathway for intracellular transport. In addition, the enhanced villi absorption in situ and oral liver distribution in vivo were also observed compared to PLGA@F127 NPs. Moreover, insulin-loaded PLGA@PPP4K NPs as an oral delivery application for diabetes induce a fine hypoglycemic response in diabetic rats after oral administration. The results of this study demonstrated that zwitterionic Pluronic analogs-coated NPs might provide a new perspective for zwitterionic materials application as well as oral delivery of biotherapeutics.

The novel role of etoposide in inhibiting the migration and proliferation of small cell lung cancer and breast cancer via targeting Daam1

ObjectivesDaam1 (Dishevelled-associated activator of morphogenesis 1) is a Wnt/PCP signaling protein that engages in cytoskeleton reorganization and is abnormally activated in certain tumors. Daam1 is closely related to cancer metastasis, which is expected to become a target for cancer treatment. However, the natural small molecules targeting Daam1 have not been identified. Materials and methodsWe screened several natural small molecules that may bind to Daam1 by Sybyl molecular simulation docking technique. As a first-line drug for the treatment of small cell lung cancer, etoposide was chosen for further investigation. Next, we used Micro Scale Thermophoresis (MST) to verify the interaction of etoposide and Daam1. Small cell lung cancer H446 cells and breast cancer MCF-7 cells were treated with etoposide and subjected to Western blotting to measure the Daam1 expression. The effect of etoposide on cell proliferation was determined by CCK-8 assay in vitro and by a tumor-bearing mouse model in vivo. Wound healing assay and Boyden chamber assay were used to evaluate the role of etoposide in the migration and invasion ability of tumor cells. The effect of etoposide on the microfilament assembly was visualized by immunofluorescence staining with phalloidine. Finally, the possible mechanism of down-regulation of Daam1 expression after etoposide-induced small cell lung cancer cells was detected by a half-life experiment and immunofluorescence staining with lysosomal marker LAMP1. ResultsSybyl molecular modeling docking technique was performed to screen a natural chemical library for molecules that bound to the FH2 domain of Daam1 and found etoposide was virtually interacted with Daam1. MST validated etoposide directly bound to the FH2 domain of Daam1. Etoposide significantly down-regulated the expression of Daam1 in small cell lung cancer H446 cells and breast cancer MCF-7 cells. Moreover, 270 μmol/L etoposide largely inhibited the proliferation, migration, and invasion of H446 cells and MCF-7 cells. Immunofluorescence staining experiments revealed that etoposide induced the disassembly of microfilaments in H446 cells and MCF-7 cells, which were rescued by Daam1 overexpression. In nude mice transplanted with H446 cells, 5, 10, 20 mg/kg etoposide (drug/weight) injected via tail vein largely retarded the proliferation of subcutaneous tumors. Etoposide induced Daam1 to shorten its half-life and enter the lysosome degradation pathway, and eventually leading to the downregulation of Daam1 expression. ConclusionsEtoposide is a novel natural small molecule targeting Daam1. Etoposide inhibits the proliferation, migration and invasion of small cell lung cancer cells and breast cancer cells, and also suppresses tumor proliferation of small cell lung cancer in vivo.

Porcine Reproductive and Respiratory Syndrome Virus nsp5 Induces Incomplete Autophagy by Impairing the Interaction of STX17 and SNAP29.

Porcine reproductive and respiratory syndrome virus (PRRSV) is an economically important pathogen that has devastated the worldwide swine industry for over 30 years. Autophagy is an evolutionarily conserved intracellular lysosomal degradation pathway, and previous studies have documented that PRRSV infection prompts autophagosome accumulation. However, whether PRRSV induces complete or incomplete autophagy remains controversial. Here, we demonstrated that overexpression of PRRSV nonstructural protein 5 (nsp5) induced the accumulation of autophagosomes, and a similar scenario was observed in PRRSV-infected cells. Moreover, both PRRSV infection and nsp5 overexpression activated incomplete autophagy, as evidenced by the blockage of autophagosome-lysosome fusion. Mechanistically, nsp5 overexpression, as well as PRRSV infection, inhibited the interaction of syntaxin 17 (STX17) with synaptosomal-associated protein 29 (SNAP29), two SNARE proteins that mediate autophagosome fusion with lysosomes, to impair the formation of autolysosomes. We further confirmed that nsp5 interacted with STX17, rather than SANP29, and the interacting domains of STX17 were the N-terminal motif and SNARE motif. Taken together, the findings of our study suggest a mechanism by which PRRSV induces incomplete autophagy by blocking autophagosome degradation and provide insights into the development of new therapeutics to combat PRRSV infection. IMPORTANCE A substantial number of viruses have been demonstrated to utilize or hijack autophagy to benefit their replication. In the case of porcine reproductive and respiratory syndrome virus (PRRSV), previous studies have demonstrated the proviral effects of autophagy on PRRSV proliferation. Thus, an investigation of the mechanism by which PRRSV regulates the autophagy processes can provide new insight into viral pathogenesis. Autophagic flux is a dynamic process that consists of autophagosome formation and subsequent lysosomal degradation. However, the exact effect of PRRSV infection on the autophagic flux remains disputed. In this study, we demonstrated that PRRSV infection, as well as PRRSV nsp5 overexpression, inhibited the interaction of STX17 with SNAP29 to impair the fusion of autophagosomes with lysosomes, thereby blocking autophagic flux. This information will help us to understand PRRSV-host interactions and unravel new targets for PRRS prevention and control.

Aptamer‐LYTACs for Targeted Degradation of Extracellular and Membrane Proteins

AbstractRecently, lysosome targeting chimeras (LYTACs) have emerged as a promising technology that expands the scope of targeted protein degradation to extracellular targets. However, the preparation of chimeras by conjugation of the antibody and trivalent N‐acetylgalactosamine (tri‐GalNAc) is a complex and time‐consuming process. The large uncertainty in number and position and the large molecular weights of the chimeras result in low internalization efficiency. To circumvent these problems, we developed the first aptamer‐based LYTAC (Apt‐LYTAC) to realize liver‐cell‐specific degradation of extracellular and membrane proteins by conjugating aptamers to tri‐GalNAc. Taking advantage of the facile synthesis and low molecular weight of the aptamer, the Apt‐LYTACs can efficiently and quickly degrade the extracellular protein PDGF and the membrane protein PTK7 through a lysosomal degradation pathway. We anticipate that the novel Apt‐LYTACs will expand the usage of aptamers and provide a new dimension for targeted protein degradation.

Aptamer-LYTACs for Targeted Degradation of Extracellular and Membrane Proteins.

Recently, lysosome targeting chimeras (LYTACs) have emerged as a promising technology that expands the scope of targeted protein degradation to extracellular targets. However, the preparation of chimeras by conjugation of the antibody and trivalent N-acetylgalactosamine (tri-GalNAc) is a complex and time-consuming process. The large uncertainty in number and position and the large molecular weights of the chimeras result in low internalization efficiency. To circumvent these problems, we developed the first aptamer-based LYTAC (Apt-LYTAC) to realize liver-cell-specific degradation of extracellular and membrane proteins by conjugating aptamers to tri-GalNAc. Taking advantage of the facile synthesis and low molecular weight of the aptamer, the Apt-LYTACs can efficiently and quickly degrade the extracellular protein PDGF and the membrane protein PTK7 through a lysosomal degradation pathway. We anticipate that the novel Apt-LYTACs will expand the usage of aptamers and provide a new dimension for targeted protein degradation.

Adipocyte autophagy limits gut inflammation by controlling oxylipin and IL‐10

Lipids play a major role in inflammatory diseases by altering inflammatory cell functions, either through their function as energy substrates or as lipid mediators such as oxylipins. Autophagy, a lysosomal degradation pathway that limits inflammation, is known to impact on lipid availability, however, whether this controls inflammation remains unexplored. We found that upon intestinal inflammation visceral adipocytes upregulate autophagy and that adipocyte‐specific loss of the autophagy gene Atg7 exacerbates inflammation. While autophagy decreased lipolytic release of free fatty acids, loss of the major lipolytic enzyme Pnpla2/Atgl in adipocytes did not alter intestinal inflammation, ruling out free fatty acids as anti‐inflammatory energy substrates. Instead, Atg7‐deficient adipose tissues exhibited an oxylipin imbalance, driven through an NRF2‐mediated upregulation of Ephx1. This shift reduced secretion of IL‐10 from adipose tissues, which was dependent on the cytochrome P450‐EPHX pathway, and lowered circulating levels of IL‐10 to exacerbate intestinal inflammation. These results suggest an underappreciated fat‐gut crosstalk through an autophagy‐dependent regulation of anti‐inflammatory oxylipins via the cytochrome P450‐EPHX pathway, indicating a protective effect of adipose tissues for distant inflammation.

Opportunities and challenges of protein-based targeted protein degradation.

In the 20 years since the first report of a proteolysis targeting chimeric (PROTAC) molecule, targeted protein degradation (TPD) technologies have attempted to revolutionize the fields of chemical biology and biomedicine by providing exciting research opportunities and potential therapeutics. However, they primarily focus on the use of small molecules to recruit the ubiquitin proteasome system to mediate target protein degradation. This then limits protein targets to cytosolic domains with accessible and suitable small molecule binding pockets. In recent years, biologics such as proteins and nucleic acids have instead been used as binders for targeting proteins, thereby expanding the scope of TPD platforms to include secreted proteins, transmembrane proteins, and soluble but highly disordered intracellular proteins. This perspective summarizes the recent TPD platforms that utilize nanobodies, antibodies, and other proteins as binding moieties to deplete challenging targets, either through the ubiquitin proteasome system or the lysosomal degradation pathway. Importantly, the perspective also highlights opportunities and remaining challenges of current protein-based TPD technologies.

Efficacy and safety clinical trial with efavirenz in patients diagnosed with adult Niemann-pick type C with cognitive impairment

Niemann-Pick disease Type C (NPC) is a genetic, incurable, neurodegenerative disorder. This orphan disease is most frequently caused by mutations in the NPC1 protein, resulting in intralysossomal cholesterol accumulation. NPC1 is found in neuronal cell bodies, axon terminals and synaptosomes, suggesting it plays a role in lysosomal degradation pathway and in synaptic transmission. Neuronal function is especially vulnerable to NPC1 deficiency and synaptic changes seem a key element in disease development. Currently, Miglustat (Zavesca®) is the only approved treatment for NPC. However, preclinical evidence showed that low-dose Efavirenz reverted synaptic defects through pharmacological activation of the enzyme CYP46. This is a single-center, phase II clinical trial to evaluate the efficacy and safety of Efavirenz in addition to standard of care in patients diagnosed with adult or late juvenile-onset NPC with cognitive impairment. All enrolled patients will be treated orally with 25 mg/d of Efavirenz for 52 weeks (1 year). Secondary objectives include evaluating clinical (neurological and neuropsychological questionnaires) and biological (imaging and biochemical biomarkers) parameters. NPC is still an unmet medical need. Although different therapeutic approaches are under study, this is the first clinical trial (to the best of our knowledge) studying the effects of Efavirenz in adult- and late-juvenile-onset NPC. Despite the small sample size and the single-arm design, we expect the results to show Efavirenz's capacity of activating the CYP46 enzyme to compensate for NPC1 deficiency and correct synaptic changes, therefore compensating cognitive and psychiatric changes in these patients. This study may provide direct benefit to enrolled patients in terms of slowing down the disease progression.

Brain-derived neurotrophic factor stimulates the retrograde pathway for axonal autophagy

Autophagy is a lysosomal degradation pathway important for neuronal development, function, and survival. How autophagy in axons is regulated by neurotrophins to impact neuronal viability and function is poorly understood. Here, we use live-cell imaging in primary neurons to investigate the regulation of axonal autophagy by the neurotrophin brain-derived neurotrophic factor (BDNF) and elucidate whether autophagosomes carry BDNF-mediated signaling information. We find that BDNF induces autophagic flux in primary neurons by stimulating the retrograde pathway for autophagy in axons. We observed an increase in autophagosome density and retrograde flux in axons, and a corresponding increase in autophagosome density in the soma. However, we find little evidence of autophagosomes comigrating with BDNF. In contrast, BDNF effectively engages its cognate receptor TrkB to undergo retrograde transport in the axon. These compartments, however, are distinct from LC3-positive autophagic organelles in the axon. Together, we find that BDNF stimulates autophagy in the axon, but retrograde autophagosomes do not appear to carry BDNF cargo. Thus, autophagosomes likely do not play a major role in relaying neurotrophic signaling information across the axon in the form of active BDNF/TrkB complexes. Rather, BDNF likely stimulates autophagy as a consequence of BDNF-induced processes that require canonical roles for autophagy in degradation.

The Emerging Role of Extracellular Vesicles and Autophagy Machinery in NASH-Future Horizons in NASH Management.

Non-alcoholic fatty liver disease (NAFLD) is considered the most frequent chronic hepatic disease in the general population, while it is the first cause of liver transplantation in the US. NAFLD patients will subsequently develop non-alcoholic steatohepatitis (NASH), which is characterized by aberrant hepatocellular inflammation with or without the presence of fibrosis. The lack of specific biomarkers and therapeutic strategies makes non-alcoholic steatohepatitis (NASH) management a difficult task for clinicians. Extracellular vesicles (EVs) constitute a heterogenic population of vesicles produced by inward or outward plasma-membrane budding. There is an emerging connection between autophagy EVs production, via an unconventional non-degradative procedure. Alterations in the amount of the secreted EVs and the cargo they carry are also involved in the disease progression and development of NASH. Autophagy constitutes a multistep lysosomal degradative pathway that reassures cell homeostasis and survival under stressful conditions, such as oxygen and energy deprivation. It prevents cellular damage by eliminating defected proteins or nοn-functional intracellular organelles. At the same time, it reassures the optimal conditions for the cells via a different mechanism that includes the removal of cargo via the secretion of EVs. Similarly, autophagy machinery is also associated with the pathogenetic mechanism of NAFLD, while it has a significant implication for the progression of the disease and the development of NASH. In this review, we will shed light on the interplay between autophagy and EVs in NASH, the emerging connection of EVs production with the autophagy pathway, and their possible manipulation for developing future therapeutic strategies for NASH.

Characterization of two novel neutral sphingomyelinase 2 inhibitors in endosomal sorting and Extracellular Vesicle biogenesis

Sphingomyelinase hydrolyzes the phosphodiester bond of the sphingomyelin to ceramide and phosphorylcholine and have been involved in extracellular vesicle (EV) biogenesis and more recently in membrane repair. Here we describe an initial testing of two recently discovered neutral sphingomyelinase 2 (nSMase2) inhibitors ((R)-(1-(3-(3,4-dimethoxyphenyl)-2,6-dimethylimidazo[1,2-b]pyridazin-8-yl)pyrrolidin-3-yl)-carbamate (PDDC) and 2,6-dimethoxy-4-[4-phenyl-5-(2-thienyl)-1H-imidazol-2-yl]phenol (DPTIP)). PDDC and DPTIP show differential effects on cell viability, and EV marker secretion, indicating that side effects of these inhibitors on lysosomal and autophagic degradation pathways need to be considered. Moreover, similar to commonly used nSMase2 inhibitor GW4869, cell type specificity seems to play a role in the endosomal trafficking routes that can be explored to unravel mechanisms of specific EV biogenesis and secretion pathways.

Macrophage autophagy in macrophage polarization, chronic inflammation and organ fibrosis.

As the essential regulators of organ fibrosis, macrophages undergo marked phenotypic and functional changes after organ injury. These changes in macrophage phenotype and function can result in maladaptive repair, causing chronic inflammation and the development of pathological fibrosis. Autophagy, a highly conserved lysosomal degradation pathway, is one of the major players to maintain the homeostasis of macrophages through clearing protein aggregates, damaged organelles, and invading pathogens. Emerging evidence has shown that macrophage autophagy plays an essential role in macrophage polarization, chronic inflammation, and organ fibrosis. Because of the high heterogeneity of macrophages in different organs, different macrophage types may play different roles in organ fibrosis. Here, we review the current understanding of the function of macrophage autophagy in macrophage polarization, chronic inflammation, and organ fibrosis in different organs, highlight the potential role of macrophage autophagy in the treatment of fibrosis. Finally, the important unresolved issues in this field are briefly discussed. A better understanding of the mechanisms that macrophage autophagy in macrophage polarization, chronic inflammation, and organ fibrosis may contribute to developing novel therapies for chronic inflammatory diseases and organ fibrosis.

Transcriptomic changes in autophagy-related genes are inversely correlated with inflammation and are associated with multiple sclerosis lesion pathology

Autophagy is a lysosomal degradative pathway essential for maintaining cellular homeostasis and is also implicated in multiple aspects of both innate and adaptive immunity. Neuroinflammation, along with demyelination and axonal loss, is an important component of multiple sclerosis (MS). Induction of autophagy ameliorated disease progression in experimental autoimmune encephalomyelitis (EAE), a mouse model for MS, underlying a possible link between autophagy and MS pathology. However, it is still unclear how autophagy is affected during different stages of MS. Here, we show a decreased expression of the autophagy-related (ATG) genes during the acute phase of EAE development in mice as well as in mixed active/inactive lesions of post-mortem human MS brain tissues. Using spatial transcriptomics, we observed that this decreased ATG gene expression is most prominent in the core of mixed active/inactive lesions. Furthermore, we observed a hyper-activation of the mammalian target of rapamycin complex 1 (mTORC1) in lesions, which could inhibit both the initiation of autophagy and the transcription factors that regulate the expression of the ATG genes. Thus, based on our data, we propose a negative regulation of autophagy in MS, possibly through persistent mTORC1 activation, which depends on the lesion stage. Our results contribute to the understanding of the role of autophagy in different stages of MS pathology and point to the mTORC1 pathway as a potential modulator that likely regulates central nervous system (CNS) homeostasis and neuroinflammation in MS.

Autophagy Dysregulation in Metabolic Associated Fatty Liver Disease: A New Therapeutic Target.

Metabolic associated fatty liver disease (MAFLD) is one of the most common causes of chronic liver disease worldwide. To date, there is no FDA-approved treatment, so there is an urgent need to determine its pathophysiology and underlying molecular mechanisms. Autophagy is a lysosomal degradation pathway that removes damaged organelles and misfolded proteins after cell injury through endoplasmic reticulum stress or starvation, which inhibits apoptosis and promotes cell survival. Recent studies have shown that autophagy plays an important role in removing lipid droplets from hepatocytes. Autophagy has also been reported to inhibit the production of pro-inflammatory cytokines and provide energy for the hepatic stellate cells activation during liver fibrosis. Thyroid hormone, irisin, melatonin, hydrogen sulfide, sulforaphane, DA-1241, vacuole membrane protein 1, nuclear factor erythroid 2-related factor 2, sodium-glucose co-transporter type-2 inhibitors, immunity-related GTPase M, and autophagy-related gene 7 have been reported to ameliorate MAFLD via autophagic induction. Lipid receptor CD36, SARS-CoV-2 Spike protein and leucine aminopeptidase 3 play a negative role in the autophagic function. This review summarizes recent advances in the role of autophagy in MAFLD. Autophagy modulates major pathological changes, including hepatic lipid metabolism, inflammation, and fibrosis, suggesting the potential of modulating autophagy for the treatment of MAFLD.

Methylation Drives SLC2A1 Transcription and Ferroptosis Process Decreasing Autophagy Pressure in Colon Cancer.

Colon cancer is a common malignant tumor in the digestive tract, with relatively high rates of morbidity and mortality. It is the third most common type of tumor in the world. The effective treatment of advanced colon cancer is limited, so it is particularly important to study the new pathogenesis of colon cancer. Ferroptosis is a nonapoptotic regulated cell death mode driven by iron-dependent lipid peroxidation, a process which has been discovered in recent years. Autophagy involves lysosomal degradation pathways that promote or prevent cell death. High levels of autophagy are associated with ferroptosis, but a clear association has not yet been made between ferroptosis and autophagy in colon cancer. Through the analysis of transcriptome expression profiling data in colon cancer, we obtained the common upregulated genes and downregulated genes by recording the intersection of the differentially expressed genes in each dataset. Solute Carrier Family 2 Member 1 (SLC2A1) was identified by combining autophagy genes obtained from GeneCards and ferroptosis genes obtained from FerrDb. In order to explore the clinical significance and prognostic value of SLC2A1, we utilized massive databases to conduct an in-depth exploration of the methylation of SLC2A1, and we also investigated the differences in immune infiltration between tumor and normal tissues. We found that there are abundant methylation sites in SLC2A1 and that the methylation of SLC2A1 is correlated with the immunosuppression of tumor tissue. We discovered that during the induction of environmental factors, the transcription and methylation levels of SLC2A1 were greatly increased, autophagy and ferroptosis were inhibited, and the immune system was defective, resulting in a poor prognosis for patients. These results suggest that the autophagy and ferroptosis-related gene SLC2A1 is involved in the tumor immune regulation of colon cancer, and SLC2A1 may become a new therapeutic target for colon cancer.

Autophagy: A Key Regulator of Homeostasis and Disease: An Overview of Molecular Mechanisms and Modulators.

Autophagy is a highly conserved lysosomal degradation pathway active at basal levels in all cells. However, under stress conditions, such as a lack of nutrients or trophic factors, it works as a survival mechanism that allows the generation of metabolic precursors for the proper functioning of the cells until the nutrients are available. Neurons, as post-mitotic cells, depend largely on autophagy to maintain cell homeostasis to get rid of damaged and/or old organelles and misfolded or aggregated proteins. Therefore, the dysfunction of this process contributes to the pathologies of many human diseases. Furthermore, autophagy is highly active during differentiation and development. In this review, we describe the current knowledge of the different pathways, molecular mechanisms, factors that induce it, and the regulation of mammalian autophagy. We also discuss its relevant role in development and disease. Finally, here we summarize several investigations demonstrating that autophagic abnormalities have been considered the underlying reasons for many human diseases, including liver disease, cardiovascular, cerebrovascular diseases, neurodegenerative diseases, neoplastic diseases, cancers, and, more recently, infectious diseases, such as SARS-CoV-2 caused COVID-19 disease.

SARS-CoV-2 spike protein inhibits megalin-mediated albumin endocytosis in proximal tubule epithelial cells

Patients with COVID-19 have high prevalence of albuminuria which is used as a marker of progression of renal disease and is associated with severe COVID-19. We hypothesized that SARS-CoV-2 spike protein (S protein) could modulate albumin handling in proximal tubule epithelial cells (PTECs) and, consequently contribute to the albuminuria observed in patients with COVID-19. In this context, the possible effect of S protein on albumin endocytosis in PTECs was investigated. Two PTEC lines were used: HEK-293A and LLC-PK1. Incubation of both cell types with S protein for 16 h inhibited albumin uptake at the same magnitude. This effect was associated with canonical megalin-mediated albumin endocytosis because: (1) DQ-albumin uptake, a marker of the lysosomal degradation pathway, was reduced at a similar level compared with fluorescein isothiocyanate (FITC)-albumin uptake; (2) dextran-FITC uptake, a marker of fluid-phase endocytosis, was not changed; (3) cell viability and proliferation were not changed. The inhibitory effect of S protein on albumin uptake was only observed when it was added at the luminal membrane, and it did not involve the ACE2/Ang II/AT1R axis. Although both cells uptake S protein, it does not seem to be required for modulation of albumin endocytosis. The mechanism underlying the inhibition of albumin uptake by S protein encompasses a decrease in megalin expression without changes in megalin trafficking and stability. These results reveal a possible mechanism to explain the albuminuria observed in patients with COVID-19.