Dysfunction of Akt/FoxO3a/Atg7 regulatory loop magnifies obesity-regulated muscular mass decline.

BACKGROUND: Myoprotein degradation accelerates in obese individuals, resulting in a decline in muscular mass. Atg7 plays a crucial role in regulating protein stability and function through both autophagy-dependent and independent pathways. As obesity progresses, the expression of Atg7 gradually rises in muscle tissue. Nonetheless, the precise impact and mechanism of Atg7 in promoting muscle mass decline in obesity remain uncertain. The study aimed to elucidate the role and underly mechanism of Atg7 action in the context of obesity-induced muscle mass decline. METHODS: In this study, we established a murine model of high-fat diet-induced obesity (DIO) and introduced adeno-associated virus delivery of short hairpin RNA to knock down Atg7 (shAtg7) into the gastrocnemius muscle. We then examined the expressions of Atg7 and myoprotein degradation markers in the gastrocnemius tissues of obese patients and mice using immunofluorescence and western blotting techniques. To further investigate the effects of Atg7, we assessed skeletal muscle cell diameter and the myoprotein degradation pathway in C2C12 and HSkMC cells in the presence or absence of Atg7. Immunofluorescence staining for MyHC and western blotting were utilized for this purpose. To understand the transcriptional regulation of Atg7 in response to myoprotein degradation, we conducted luciferase reporter assays and chromatin immunoprecipitation experiments to examine whether FoxO3a enhances the transcription of Atg7. Moreover, we explored the role of Akt in Atg7-mediated regulation and its relevance to obesity-induced muscle mass decline. This was accomplished by Akt knockdown, treatment with MK2206, and GST pulldown assays to assess the interaction between Atg7 and Akt. RESULTS: After 20 weeks of being on a high-fat diet, obesity was induced, leading to a significant decrease in the gastrocnemius muscle area and a decline in muscle performance. This was accompanied by a notable increase in Atg7 protein expression (p < 0.01). Similarly, in gastrocnemius tissues of obese patients when compared to nonobese individuals, there was a significant increase in both Atg7 (p < 0.01) and TRIM63 (p < 0.01) levels. When palmitic acid was administered to C2C12 cells, it resulted in increased Atg7 (p < 0.01), LC3Ⅱ/Ⅰ (p < 0.01), and p62 levels (p < 0.01). Additionally, it promoted FoxO3a-mediated transcription of Atg7. The knockdown of Atg7 in the gastrocnemius partially reversed DIO-induced muscle mass decline. Furthermore, when Atg7 was knocked down in C2C12 and HSkMC cells, it mitigated palmitic acid-induced insulin resistance, increased the p-Akt/Akt ratio (p < 0.01), and reduced TRIM63 (p < 0.01). Muscular atrophy mediated by Atg7 was reversed by genetic knockdown of Akt and treatment with the p-Akt inhibitor MK2206. Palmitic acid administration increased the binding between Atg7 and Akt (p < 0.01) while weakening the binding of PDK1 (p < 0.01) and PDK2 (p < 0.01) to Akt. GST pulldown assays demonstrated that Atg7 directly interacted with the C-terminal domain of Akt. CONCLUSION: The consumption of a high-fat diet, along with lipid-induced effects, led to the inhibition of Akt signaling, which, in turn, promoted FoxO3a-mediated transcription, increasing Atg7 levels in muscle cells. The excess Atg7 inhibited the phosphorylation of Akt, leading to a cyclic activation of FoxO3a and exacerbating the decline in muscle mass regulated by obesity. Consequently, Atg7 serves as a regulatory point in determining the decline in muscle mass induced by obesity.

Adenovirus-mediated Overexpression of FcγRIIB Attenuates Pulmonary Inflammation and Fibrosis.

Progressive fibrosing interstitial lung diseases (PF-ILDs) result in high mortality and lack effective therapies. The pathogenesis of PF-ILDs involves macrophages driving inflammation and irreversible fibrosis. Fc-γ receptors (FcγRs) regulate macrophages and inflammation, but their roles in PF-ILDs remain unclear. We characterized the expression of FcγRs and found upregulated FcγRIIB in human and mouse lungs after exposure to silica. FcγRIIB deficiency aggravated lung dysfunction, inflammation, and fibrosis in silica-exposed mice. Using single-cell transcriptomics and in vitro experiments, FcγRIIB was found in alveolar macrophages, where it regulated the expression of fibrosis-related genes Spp1 and Ctss. In mice with macrophage-specific overexpression of FcγRIIB and in mice treated with adenovirus by intratracheal instillation to upregulate FcγRIIB, silica-induced functional and histological changes were ameliorated. Our data from three genetic models and a therapeutic model suggest that FcγRIIB plays a protective role that can be enhanced by adenoviral overexpression, representing a potential therapeutic strategy for PF-ILDs.

FcεRI deficiency alleviates silica-induced pulmonary inflammation and fibrosis.

Silicosis is one of the most important occupational diseases worldwide, caused by inhalation of silica particles or free crystalline silicon dioxide. As a disease with high mortality, it has no effective treatment and new therapeutic targets are urgently needed. Recent studies have identified FCER1A, encoding α-subunit of the immunoglobulin E (IgE) receptor FcεRI, as a candidate gene involved in the biological pathways leading to respiratory symptoms. FcεRI is known to be important in allergic asthma, but its role in silicosis remains unclear. In this study, serum IgE concentrations and FcεRI expression were assessed in pneumoconiosis patients and silica-exposed mice. The role of FcεRI was explored in a silica-induced mouse model using wild-type and FcεRI-deficient mice. The results showed that serum IgE concentrations were significantly elevated in both pneumoconiosis patients and mice exposed to silica compared with controls. The mRNA and protein expression of FcεRI were also significantly increased in the lung tissue of patients and silica-exposed mice. FcεRI deficiency significantly attenuated the changes in lung function caused by silica exposure. Silica-induced elevations of IL-1ß, IL-6, and TNF-α were significantly attenuated in the lung tissue and bronchoalveolar lavage fluid (BALF) of FcεRI-deficient mice compared with wild-type controls. Additionally, FcεRI-deficient mice showed a significantly lower score of pulmonary fibrosis than wild-type mice following exposure to silica, with significantly lower hydroxyproline content and expression of fibrotic genes Col1a1 and Fn1. Immunofluorescent staining suggested FcεRI mainly on mast cells. Mast cell degranulation took place after silica exposure, as shown by increased serum histamine levels and ß-hexosaminidase activity, which were significantly reduced in FcεRI-deficient mice compared with wild-type controls. Together, these data showed that FcεRI deficiency had a significant protective effect against silica-induced pulmonary inflammation and fibrosis. Our findings provide new insights into the pathophysiological mechanisms of silica-induced pulmonary fibrosis and a potential target for the treatment of silicosis.

Nur77 ameliorates age-related renal tubulointerstitial fibrosis by suppressing the TGF-ß/Smads signaling pathway.

Nerve growth factor-induced gene B (Nur77) has been shown to ameliorate several biological processes in chronic diseases, including inflammatory response, cellular proliferation, and metabolism. Chronic kidney disease (CKD) is characterized by tubulointerstitial fibrosis for which no targeted therapies are available as yet. In this study, we performed in vivo and in vitro experiments to demonstrate that Nur77 targets fibrosis signals and attenuates renal tubulointerstitial fibrosis during the aging process. We observed that the TGF-ß/Smads signal pathway was significantly suppressed by Nur77, suggesting that Nur77 controlled the activation of key steps in TGF-ß/Smads signaling. We further showed that Nur77 interacted with Smad7, the main repressor of nuclear translocation of Smad2/3, and stabilized Smad7 protein homeostasis. Nur77 deficiency resulted in Smad7 degradation, aggravating Smad2/3 phosphorylation, and promoting transcription of its downstream target genes, ACTA2 and collagen I. Our findings demonstrate that Nur77 is a potential therapeutic target for age-related kidney diseases including CKD. Maintenance of Nur77 may be an effective strategy for blocking renal tubulointerstitial fibrosis and improving renal function in the elderly.

Berberine modulates deacetylation of PPARγ to promote adipose tissue remodeling and thermogenesis via AMPK/SIRT1 pathway.

Pharmacological stimulation of adipose tissue remodeling and thermogenesis to increase energy expenditure is expected to be a viable therapeutic strategy for obesity. Berberine has been reported to have pharmacological activity in adipose tissue to anti-obesity, while the mechanism remains unclear. Here, we observed that berberine significantly reduced the body weight and insulin resistance of high-fat diet mice by promoting the distribution of brown adipose tissue and thermogenesis. We have further demonstrated that berberine activated energy metabolic sensing pathway AMPK/SIRT1 axis to increase the level of PPARγ deacetylation, which leads to promoting adipose tissue remodeling and increasing the expression of the thermogenic protein UCP-1. These findings suggest that berberine that enhances the AMPK/SIRT1 pathway can act as a selective PPARγ activator to promote adipose tissue remodeling and thermogenesis. This study proposes a new mechanism for the regulation of berberine in adipose tissue and offers a great prospect for berberine in obesity treatment.

Characteristics of Online Health Care Services From China's Largest Online Medical Platform: Cross-sectional Survey Study.

BACKGROUND: Internet hospitals in China are in great demand due to limited and unevenly distributed health care resources, lack of family doctors, increased burdens of chronic diseases, and rapid growth of the aged population. The COVID-19 epidemic catalyzed the expansion of online health care services. In recent years, internet hospitals have been rapidly developed. Ping An Good Doctor is the largest, national online medical entry point in China and is a widely used platform providing online health care services. OBJECTIVE: This study aims to give a comprehensive description of the characteristics of the online consultations and inquisitions in Ping An Good Doctor. The analyses tried to answer the following questions: (1) What are the characteristics of the consultations in Ping An Good Doctor in terms of department and disease profiles? (2) Who uses the online health services most frequently? and (3) How is the user experience of the online consultations of Ping An Good Doctor? METHODS: A total of 35.3 million consultations and inquisitions over the course of 1 year were analyzed with respect to the distributions of departments and diseases, user profiles, and consulting behaviors. RESULTS: The geographical distribution of the usage of Ping An Good Doctor showed that Shandong (18.4%), Yunnan (15.6%), Shaanxi (7.2%), and Guangdong (5.5%) were the provinces that used it the most; they accounted for 46.6% of the total consultations and inquisitions. In terms of department distribution, we found that gynecology and obstetrics (19.2%), dermatology (17.0%), and pediatrics (14.4%) were the top three departments in Ping An Good Doctor. The disease distribution analysis showed that, except for nondisease-specific consultations, acute upper respiratory infection (AURI) (4.1%), pregnancy (2.8%), and dermatitis (2.4%) were the most frequently consulted diseases. In terms of user profiles, females (60.4%) from 19 to 35 years of age were most likely to seek consultations online, in general. The user behavior analyses showed that the peak times of day for online consultations occurred at 10 AM, 3 PM, and 9 PM. Regarding user experience, 93.0% of users gave full marks following their consultations. For some disease-related health problems, such as AURI, dermatitis, and eczema, the feedback scores were above average. CONCLUSIONS: The prevalence of internet hospitals, such as Ping An Good Doctor, illustrated the great demand for online health care services that can go beyond geographical limitations. Our analyses showed that nondisease-specific issues and moderate health problems were much more frequently consulted about than severe clinical conditions. This indicated that internet hospitals played the role of the family doctor, which helped to relieve the stress placed on offline hospitals and facilitated people's lives. In addition, good user experiences, especially regarding disease-related inquisitions, suggested that online health services can help solve health problems. With support from the government and acceptance by the public, online health care services could develop at a fast pace and greatly benefit people's daily lives.

Construction of a Non-Mutually Exclusive Decision Tree for Medication Recommendation of Chronic Heart Failure.

Objective: Although guidelines have recommended standardized drug treatment for heart failure (HF), there are still many challenges in making the correct clinical decisions due to the complicated clinical situations of HF patients. Each patient would satisfy several recommendations, meaning the decision tree of HF treatment should be nonmutually exclusive, and the same patient would be allocated to several leaf nodes in the decision tree. In the current study, we aim to propose a way to ensemble a nonmutually exclusive decision tree for recommendation system for complicated diseases, such as HF. Methods: The nonmutually exclusive decision tree was constructed via knowledge rules summarized from the HF clinical guidelines. Then similar patients were defined as those who followed the same pattern of leaf node allocation according to the decision tree. The frequent medication patterns for each similar patient were mined using the Apriori algorithms, and we also carried out the outcome prognosis analyses to show the capability for the evidence-based medication recommendations of our nonmutually exclusive decision tree. Results: Based on a large database that included 29,689 patients with 84,705 admissions, we tested the framework for HF treatment recommendation. In the constructed decision tree, the HF treatment recommendations were grouped into two independent parts. The first part was recommendations for new cases, and the second part was recommendations when patients had different historical medication. There are 14 leaf nodes in our decision tree, and most of the leaf nodes had a guideline adherence of around 90%. We reported the top 10 popular similar patients, which accounted for 32.84% of the whole population. In addition, the multiple outcome prognosis analyses were carried out to assess the medications for one of the subgroups of similar patients. Our results showed even for the subgroup of the same similar patients that no one medication pattern would benefit all outcomes. Conclusion: In the present study, the methodology to construct a nonmutually exclusive decision tree for medication recommendations for HF and its application in CDSS was proposed. Our framework is universal for most diseases and could be generally applied in developing the CDSS for treatment.

Pharmacological Activation of Estrogen Receptor Beta Overcomes Tumor Resistance to Immune Checkpoint Blockade Therapy.

The emerging immune checkpoint blockade (ICB) therapy has ushered the cancer therapeutics field into an era of immunotherapy. Although ICB treatment provides remarkable clinical responses in a subset of patients with cancer, this regimen fails to extend survival in a large proportion of patients. Here, we found that a combined treatment of estrogen receptor beta (ERß) agonist and PD-1 antibody treatment improved therapeutic efficacy in mouse tumor models, compared with monotherapies, by reducing infiltration of myeloid-derived suppressor cells (MDSCs) and increasing CD8+ T cells in tumors. Mechanistically, LY500307 treatment reduced tumor-derived CSF1 and decreased infiltration of CSF1R+ MDSCs in the tumor bed. CSF1 released by tumor cells induced CSF1R+ MDSC chemotaxis in vitro and blockade of CSF1R demonstrated similar therapeutic effects as ERß activation in vivo. Collectively, our study proved combined treatment of ERß agonist and PD-1 antibody reduced MDSC infiltration in the tumor and enhanced tumor response to ICB therapy.

Construction of Guideline-Based Decision Tree for Medication Recommendation.

Clinical decision support system (CDSS) plays an essential role nowadays and CDSS for treatment provides clinicians with the clinical evidence of candidate prescriptions to assist them in making patient-specific decisions. Therefore, it is essential to find a partition of patients such that patients with similar clinical conditions are grouped together and the preferred prescriptions for different groups are diverged. A comprehensive clinical guideline often provides information of patient partition. However, for most diseases, the guideline is not so detailed that only limited circumstances are covered. This makes it challenging to group patients properly. Here we proposed an approach that combines clinical guidelines with medical data to construct a nested decision tree for patient partitioning and treatment recommendation. Compared with pure data-driven decision tree, the recommendations generated by our model have better guideline adherence and interpretability. The approach was successfully applied in a real-world case study of patients with hyperthyroidism.

α-Helical Motif as Inhibitors of Toxic Amyloid-ß Oligomer Generation via Highly Specific Recognition of Amyloid Surface.

Amyloid fibril surfaces can convert soluble proteins into toxic oligomers and are attractive targets for intervention of protein aggregation diseases. Thus far, molecules identified with inhibitory activity are either large proteins or flat cyclic compounds lacking in specificity. The main design difficulty is flatness of amyloid surfaces and the lack of knowledge on binding interfaces. Here, we demonstrate, for the first time, a rational design of alpha-helical peptide inhibitors targeting the amyloid-beta 40 (Aß40) fibril surfaces, based on our in silico finding that a helical fragment of Aß40 interacts in a unique way with side-chain arrays on the fibril surface. We strengthen the fragment's binding capability through mutations and helicity enhancement with our Terminal Aspartic acid strategy. The resulting inhibitor shows micromolar affinity for the fibril surface, effectively impedes the surface-mediated oligomerization of Aß40, and mitigates its cytotoxicity. This work opens up an avenue to designing aggregation modulators for amyloid diseases.

Integrating Clinical Knowledge and Real-World Evidence for Type 2 Diabetes Treatment.

Clinical decision support system (CDSS) plays a significant role nowadays and it assists physicians in making decisions for treatment. Generally based on clinical guideline, the principles of the recommendation are provided and may suggest several candidate medications for similar patient group with certain clinical conditions. However, it is challenging to prioritize these candidates and even refine the guideline to a finer level for patient-specific recommendation. Here we propose a method and system to integrate the clinical knowledge and real-world evidence (RWE) for type 2 diabetes treatment, to enable both standardized and personalized medication recommendation. The RWE is generated by medication effectiveness analysis and subgroup analysis. The knowledge model has been verified by clinical experts from the advanced hospitals. The data verification results show that the medications that are consistent with the method recommendation can lead to better clinical outcome in terms of glycemic control, compared to those inconsistent.

In Silico Study of Recognition between Aß40 and Aß40 Fibril Surfaces: An N-Terminal Helical Recognition Motif and Its Implications for Inhibitor Design.

The recent finding that the surface of amyloid-ß (Aß) fibril can recruit Aß peptides and convert them into toxic oligomers has rendered fibril surfaces attractive as inhibition targets. Through extensive simulations with hybrid-resolution and all-atom models, we have investigated how Aß1-40 recognizes its own fibril surfaces. These calculations give a ∼2.6-5.6 µM half-saturation concentration of Aß on the surface (cf. experimental value ∼6 µM). Aß was found to preferentially bind to region 16-24 of Aß40 fibrils through both electrostatic and van der Waals forces. Both terminal regions of Aß contribute significantly to binding energetics. A helical binding pose of the N-terminal region of Aß (Aß3-14) not seen before is highly preferred on the fibril surface. Aß3-14 in a helical form can arrange side chains with similar properties on the same sides of the helix and maximize complementary interactions with side chain arrays characteristic of amyloid fibrils. Helix formation on a fibril surface implies a helix-mediated mechanism for Aß oligomerization catalyzed by fibrils. We propose an Aß3-14 analogue that can exhibit enhanced helical character and interactions with Aß fibrils and may thus be used as a template with which to pursue potent inhibitors of Aß-fibril interactions.

Self-Assembly Pathways of ß-Sheet-Rich Amyloid-ß(1-40) Dimers: Markov State Model Analysis on Millisecond Hybrid-Resolution Simulations.

Early oligomerization during amyloid-ß (Aß) aggregation is essential for Aß neurotoxicity. Understanding how unstructured Aßs assemble into oligomers, especially those rich in ß-sheets, is essential but remains challenging as the assembly process is too transient for experimental characterization and too slow for molecular dynamics simulations. So far, atomic simulations are limited only to studies of either oligomer structures or assembly pathways for short Aß segments. To overcome the computational challenge, we combine in this study a hybrid-resolution model and adaptive sampling techniques to perform over 2.7 ms of simulations of formation of full-length Aß40 dimers that are the earliest toxic oligomeric species. The Markov state model is further employed to characterize the transition pathways and associated kinetics. Our results show that for two major forms of ß-sheet-rich structures reported experimentally, the corresponding assembly mechanisms are markedly different. Hairpin-containing structures are formed by direct binding of soluble Aß in ß-hairpin-like conformations. Formation of parallel, in-register structures resembling fibrils occurs ∼100-fold more slowly and involves a rapid encounter of Aß in arbitrary conformations followed by a slow structural conversion. The structural conversion proceeds via diverse pathways but always requires transient unfolding of encounter complexes. We find that the transition kinetics could be affected differently by intra-/intermolecular interactions involving individual residues in a conformation-dependent manner. In particular, the interactions involving Aß's N-terminal part promote the assembly into hairpin-containing structures but delay the formation of fibril-like structures, thus explaining puzzling observations reported previously regarding the roles of this region in the early assembly process.