Differential Clearance of Aβ Species from the Brain by Brain Lymphatic Endothelial Cells in Zebrafish

The failure of amyloid beta (Aβ) clearance is a major cause of Alzheimer’s disease, and the brain lymphatic systems play a crucial role in clearing toxic proteins. Recently, brain lymphatic endothelial cells (BLECs), a non-lumenized lymphatic cell in the vertebrate brain, was identified, but Aβ clearance via this novel cell is not fully understood. We established an in vivo zebrafish model using fluorescently labeled Aβ42 to investigate the role of BLECs in Aβ clearance. We discovered the efficient clearance of monomeric Aβ42 (mAβ42) compared to oligomeric Aβ42 (oAβ42), which was illustrated by the selective uptake of mAβ42 by BLECs and peripheral transport. The genetic depletion, pharmacological inhibition via the blocking of the mannose receptor, or the laser ablation of BLECs resulted in the defective clearance of mAβ42. The treatment with an Aβ disaggregating agent facilitated the internalization of oAβ42 into BLECs and improved the peripheral transport. Our findings reveal a new role of BLECs in the differential clearance of mAβ42 from the brain and provide a novel therapeutic strategy based on promoting Aβ clearance.

KLF15 Regulates Oxidative Stress Response in Cardiomyocytes through NAD+

KLF15 has recently emerged as a central regulator of metabolism. Although its connection to oxidative stress has been suspected, there has not been any study to date that directly demonstrates the molecular link. In this study, we sought to determine the role of KLF15 in cardiac oxidative stress. We found that KLF15 deficiency in the heart is associated with increased oxidative stress. Acute deficiency of KLF15 in neonatal rat ventricular myocytes (NRVMs) leads to the defective clearance of reactive oxygen species (ROS) and an exaggerated cell death following a variety of oxidative stresses. Mechanistically, we found that KLF15 deficiency leads to reduced amounts of the rate-limiting NAD+ salvage enzyme NAMPT and to NAD+ deficiency. The resultant SIRT3-dependent hyperacetylation and the inactivation of mitochondrial antioxidants can be rescued by MnSOD mimetics or NAD+ precursors. Collectively, these findings suggest that KLF15 regulates cardiac ROS clearance through the regulation of NAD+ levels. Our findings establish KLF15 as a central coordinator of cardiac metabolism and ROS clearance.

Reduced astrocytic reactivity in human brains and midbrain organoids with PRKN mutations

Parkin (encoded by PRKN) is a ubiquitin ligase that plays an important role in cellular mitochondrial quality control. Mutations in PRKN cause selective dopaminergic cell loss in the substantia nigra and are presumed to induce a decrease in mitochondrial function caused by the defective clearance of mitochondria. Several studies have demonstrated that parkin dysfunction causes mitochondrial injury and astrocytic dysfunction. Using immunohistochemical methods, we analyzed astrocytic changes in human brains from individuals with PRKN mutations. Few glial fibrillary acidic protein- and vimentin-positive astrocytes were observed in the substantia nigra in PRKN-mutated subjects compared with subjects with idiopathic Parkinson’s disease. We also differentiated patient-specific induced pluripotent stem cells into midbrain organoids and confirmed decreased numbers of glial fibrillary acidic protein-positive astrocytes in PRKN-mutated organoids compared with age- and sex-matched controls. Our study reveals PRKN-mutation-induced astrocytic alteration and suggests the possibility of an astrocyte-related non-autonomous cell death mechanism for dopaminergic neurons in brains of PRKN-mutated patients.

Bcl-2-associated athanogene 5 (BAG5) regulates Parkin-dependent mitophagy and cell death

As pathogenic Parkin mutations result in the defective clearance of damaged mitochondria, Parkin-dependent mitophagy is thought to be protective against the dopaminergic neurodegeneration observed in Parkinson’s disease. Recent studies, however, have demonstrated that Parkin can promote cell death in the context of severe mitochondrial damage by degrading the pro-survival Bcl-2 family member, Mcl-1. Therefore, Parkin may act as a ‘switch’ that can shift the balance between protective or pro-death pathways depending on the degree of mitochondrial damage. Here, we report that the Parkin interacting protein, Bcl-2-associated athanogene 5 (BAG5), impairs mitophagy by suppressing Parkin recruitment to damaged mitochondria and reducing the movement of damaged mitochondria into the lysosomes. BAG5 also enhanced Parkin-mediated Mcl-1 degradation and cell death following severe mitochondrial insult. These results suggest that BAG5 may regulate the bi-modal activity of Parkin, promoting cell death by suppressing Parkin-dependent mitophagy and enhancing Parkin-mediated Mcl-1 degradation.

ANTIHYPERLIPIDEMIC ACTIVITY OF METHANOLIC EXTRACT OF AZADIRACHTA INDICA

Hyperlipidemia often results from delayed or defective clearance, or overproduction of VLDL by the liver, which is subsequently transformed into LDL. The whole plant of Azadirachta indica was collected from local areas of Jaipur district areas of Rajasthan. The plant is authenticated by university of rajasthan by Drmanju Sharma dept of botany Selected medicinal plant were cut into small pieces, cleaned and shade dried at room temperature then subjected to physical evaluation with different parameters. Macerated 5 g of air dried drug coarsely powdered with 100 ml of ethanol of the specified strength in a closed flask for twenty four hours, shaking frequently during six hours and allowed to stand for 18 h. About 2-3 g of powder is accurately weighed in a china dish and kept in hot air oven maintained at 110±1oC for four hours. After cooling in a desiccator, the loss in weight was recorded. This procedure was repeated till constant weight was obtained. For insoluble ash value Boiled the ash for 5-10 minutes with 25 ml of diluted hydrochloric acid, collected the insoluble matter in a Gooch crucible washed with hot water, ignited and weighed. Whole plant of Azadirachtaindicawas reduced to fine powder and was subjected to maceration in a 1 liter conical flask for 7 days at room temperature with alcohol (70%) water (30%). In the present study we aimed to screen various extracts of selected plants on Cholesterol induced hyperlipidemic rat model. Thus the results of the present investigation clearly indicated that the selected medicinal plants possess good antihyerlipidemic activity in atherogenic diet induced hyperlipidemic rats and led to the development of new Herbal formulation possessing antihyperlipidemic and antiatherosclerotic activities. This is the first study which investigates the hypolipidemic activity. Keywords: Azadirachta indica, antihyperlipidemic, antiatherosclerotic activities, Maceration.

Molecular and Metabolic Pathogenesis of Familial Combined Hyperlipidemia and Association with Metabolic Syndrome

Background The objective of this review is to unify the various genetic defects along with elaborating metabolic pathways in Familial Combined Hyperlipidemia(FCHL) and also to differentiate the phenotype of FCHL from metabolic syndrome. Methods PubMed and Cochrane’s library was searched for keyword “Familial combined hyperlipidemia” and latter with “Familial combined hyperlipidemia genes” to finally shortlist 23 articles. Further search with key words “molecular pathogenesis of familial combined hyperlipidemia” and “metabolic syndrome and familial combined hyperlipidemia” was carried out for finding molecular defects in FCHL, non-molecular findings distinguishing FCHL from metabolic syndrome and overlapping features between FCHL and metabolic syndrome. Results Major culprit regions identified included Chromosome-1q21-q24(USF1 and FOXA2) , Ch-11q (APOA5), Ch-16q24, Ch-20q12-q13.1, Ch.4q32.3 (rs6829588), and Ch-19q13.32 containing PVRL-2 gene (Also known as Nectin-2). The genetic and metabolic pathways linked to FCHL may involve: 1-Defective clearance of Apo-B containing lipoproteins, 2-Overproduction of Apo-B containing lipoprotein i.e., VLDL and 3-Adipose tissue dysfunction. FCHL phenotype showed close resemblance with metabolic syndrome clinical and biochemical features with slight differences. Conclusion The reviewed data suggested that FCHL phenotype is the resultant end outcome from multiple molecular defects and thus underlying genetic defect identification in the index case is important for personalized medicine and incoming gene therapy. Further research is warranted to explore specific genetic defects.

PERAN IMUNITAS HUMORAL PADA PENYAKIT SYSTEMIC LUPUS ERYTHEMATOSUS (SLE)

Systemic Lupus Erythematosus (SLE) adalah suatu kelainan autoimun multisistem kronik yang ditandai oleh autoantibodi dan kompleks imun yang berkaitan dengan manifestasi klinis dan kerusakan jaringan yang beragam dan luas. Beberapa defek dari komponen imunologi multipel berperan penting dalam patogenesis SLE. Abnormalitas imunologi pada SLE meliputi kemampuan untuk menghasilkan autoantibodi patogenik, hilangnya regulasi sel limfosit B dan T, serta defective clearance dari autoantigen dan kompleks imun. Faktor genetik, imunologik dan hormonal serta lingkungan diduga berperan dalam patofisiologi SLE. Tulisan ini mengkaji respon imun humoral sebagai salah satu patogenesis SLE.

Dual Role for CIB1 in Thrombopoiesis: CIB1 Suppresses Megakaryocyte Quantities but Supports Adhesion, Migration, and Proplatelet Formation

Abstract 2384 Megakaryocytes (MKs) are large polyploid cells that produce platelets through a process known as thrombopoiesis. Thrombopoietin (Tpo) is the major cytokine that regulates a variety of steps in this process, including hematopoietic stem cell (HSC) differentiation to MKs, proplatelet formation, and platelet release into the circulation. However, the molecular mechanism of thrombopoiesis is poorly understood. We have previously reported that calcium- and integrin-binding protein 1 (CIB1) regulates endomitosis in Dami cells. To further characterize the role of CIB1 in thrombopoiesis, we utilized a Cib1−/− mouse model. We observed that Cib1−/− mice have a slightly elevated number of platelets and bone marrow (BM)-derived MKs than wild-type (WT) controls (p<0.05). Rate of platelet clearance was comparable in Cib1−/− and WT mice, suggesting that the defective clearance is not the cause of the observed elevated platelet number. In order to determine if the HSC differentiation is dysregulated by the ablation of Cib1, we analyzed MK-colony forming unit production, which revealed an increase in the colony forming cells with Cib1 deletion compared to WT (p<0.05). Additionally, BM from Cib1−/− mice, cultured with Tpo for 24 hours, produced more highly polyploid MKs than WT BM (p<0.05). These results suggest that Cib1 may negatively regulate initial steps of megakaryopoiesis. Subsequent analysis of Tpo signaling revealed that activation of FAK, a known suppresser of Tpo signaling, is attenuated, as indicated by reduced FAKY925 phosphorylation in Cib1−/− BM-derived MKs treated with Tpo. Consequently, Akt and ERK1/2 activation downstream of Tpo was enhanced. These results suggested that Cib1 inhibits Tpo signaling by augmenting FAK activation. Interestingly, platelet recovery in Cib1−/− mice following platelet depletion by experimental immunothrombocytopenia was attenuated compared to WT (p<0.05). This could be due to impaired adhesion and migration of MKs on the extracellular matrix. Consistent with this notion, adhesion to fibrinogen and fibronectin and migration towards an SDF-1α gradient were significantly reduced in Cib1−/− MKs compared to WT (p<0.05). Additionally, Cib1−/− MKs formed fewer proplatelets compared to WT (p<0.05), when plated on fibrinogen. These data suggest that CIB1 plays a dual role in thrombopoiesis, initially by negatively regulating Tpo signaling, and later by supporting MK migration and proplatelet production. Disclosures: No relevant conflicts of interest to declare.

Regulation of steady-state neutrophil homeostasis by macrophages

The timely clearance of apoptotic neutrophils from inflammation sites is an important function of macrophages; however, the role of macrophages in maintaining neutrophil homeostasis under steady-state conditions is less well understood. By conditionally deleting the antiapoptotic gene cellular FLICE-like inhibitory protein (C-FLIP) in myeloid cells, we have generated a novel mouse model deficient in marginal zone and bone marrow stromal macrophages. These mice develop severe neutrophilia, splenomegaly, extramedullary hematopoiesis, decreased body weight, and increased production of granulocyte colony-stimulating factor (G-CSF) and IL-1β, but not IL-17. c-FLIPf/f LysM-Cre mice exhibit delayed clearance of circulating neutrophils, suggesting that failure of macrophages to efficiently clear apoptotic neutrophils causes production of cytokines that drive excess granulopoiesis. Further, blocking G-CSF but not IL-1R signaling in vivo rescues this neutrophilia, suggesting that a G-CSF–dependent, IL-1β–independent pathway plays a role in promoting neutrophil production in mice with defective clearance of apoptotic cells.