Disease severity–related alterations of cardiac microRNAs in experimental pulmonary hypertension

Right ventricular (RV) failure is the primary cause of death in pulmonary arterial hypertension (PAH). We hypothesized that heart‐relevant microRNAs, that is myomiRs (miR‐1, miR‐133a, miR‐208, miR‐499) and miR‐214, can have a role in the right ventricle in the development of PAH. To mimic PAH, male Wistar rats were injected with monocrotaline (MCT, 60 mg/kg, s.c.); control group received vehicle. MCT rats were divided into two groups, based on the clinical presentation: MCT group terminated 4 weeks after MCT administration and prematurely terminated group (ptMCT) displaying signs of terminal disease. Myocardial damage genes and candidate microRNAs expressions were determined by RT‐qPCR. Reduced blood oxygen saturation, breathing disturbances, RV enlargement as well as elevated levels of markers of myocardial damage confirmed PH in MCT animals and were more pronounced in ptMCT. MyomiRs (miR‐1/miR‐133a/miR‐208a/miR‐499) were decreased and the expression of miR‐214 was increased only in ptMCT group (P < 0.05). The myomiRs negatively correlated with Fulton index as a measure of RV hypertrophy in MCT group (P < 0.05), whereas miR‐214 showed a positive correlation (P < 0.05). We conclude that the expression of determined microRNAs mirrored the disease severity and targeting their pathways might represent potential future therapeutic approach in PAH.     

Inhibition of neuronal necroptosis mediated by RIP1/RIP3/MLKL provides neuroprotective effects on kaolin‐induced hydrocephalus in mice

ObjectivesNecroptosis is widespread in neurodegenerative diseases. Here, we examined necroptosis in the hippocampus and cortex after hydrocephalus and found that a necroptosis pathway inhibitor alleviates necroptosis and provides neuroprotective effects.Materials and methodsHydrocephalus was induced in C57BL/6 mice by kaolin. Haematoxylin and eosin (HE), Nissl, PI and Fluoro‐Jade B (FJB) staining were used for general observations. Phosphorylated receptor‐interacting protein kinase 3 (p‐RIP3) and phosphorylated mixed lineage kinase domain‐like (p‐MLKL) were measured by Western blotting and immunohistochemistry. Scanning electron microscopy (SEM) was used to observe ependymal cilia. Magnetic resonance imaging (MRI) and the Morris water maze (MWM) test were used to assess neurobehavioral changes. Immunofluorescence was used to detect microglial and astrocyte activation. Inflammatory cytokines were measured by Western blotting and RT‐PCR.ResultsObvious pathological changes appeared in the hippocampus and cortex after hydrocephalus, and expression of the necroptosis markers p‐RIP3, p‐MLKL and inflammatory cytokines increased. Necrostatin‐1 (Nec‐1) and GSK872 reduced necrotic cell death, attenuated p‐RIP3 and p‐MLKL levels, slightly improved neurobehaviours and inhibited microglial and astrocyte activation and inflammation.ConclusionsRIP1/RIP3/MLKL mediates necroptosis in the cortex and hippocampus in a hydrocephalus mouse model, and Nec‐1 and GSK872 have some neuroprotective effects.     

MiR‐223‐3p inhibits rTp17‐induced inflammasome activation and pyroptosis by targeting NLRP3

The incidence of syphilis caused by Treponema pallidum subsp pallidum (T pallidum) infection is accompanied by inflammatory injuries of vascular endothelial cells. Studies have revealed that T pallidum infection could induce inflammasome activation and pyroptosis in macrophages. MicroRNA‐223‐3p (miR‐223‐3p) was reported to be a negative regulator in inflammatory diseases. The present study aimed to explore whether miR‐223‐3p regulates T pallidum‐induced inflammasome activation and pyroptosis in vascular endothelial cells, and determine the mechanisms which underlie this process. MiR‐223‐3p levels in syphilis and control samples were determined. The biological function of miR‐223‐3p in the NLRP3 inflammasome and pyroptosis was evaluated in T pallidum‐infected human umbilical vein endothelial cells (HUVECs). We observed a dramatic decrease in miR‐223‐3p levels in syphilis patients (n = 20) when compared to healthy controls (n = 20). Moreover, miR‐223‐3p showed a notable inhibitory effect on recombinant Tp17 (rTP17)‐induced caspase‐1 activation, resulting in decrease in IL‐1β production and pyroptosis, which was accompanied by the release of lactate dehydrogenase (LDH) in HUVECs. Additionally, the dual‐luciferase assay confirmed that NLRP3 is a direct target of miR‐223‐3p. Moreover, NLRP3 overexpression or knockdown largely blocked the effects of miR‐223‐3p on T pallidum‐induced inflammasome activation and pyroptosis in HUVECs. Most importantly, a notable negative correlation was observed between miR‐223‐3p and NLRP3, caspase‐1, and IL‐1β, respectively, in the serum of syphilis patients and healthy controls. Taken together, our results reveal that miR‐223‐3p targets NLRP3 to suppress inflammasome activation and pyroptosis in T pallidum‐infected endothelial cells, implying that miR‐223‐3p could be a potential target for syphilis patients.     

Necroptosis contributes to chronic inflammation and fibrosis in aging liver

Inflammaging, characterized by an increase in low‐grade chronic inflammation with age, is a hallmark of aging and is strongly associated with various age‐related diseases, including chronic liver disease (CLD) and hepatocellular carcinoma (HCC). Because necroptosis is a cell death pathway that induces inflammation through the release of DAMPs, we tested the hypothesis that age‐associated increase in necroptosis contributes to chronic inflammation in aging liver. Phosphorylation of MLKL and MLKL oligomers, markers of necroptosis, as well as phosphorylation of RIPK3 and RIPK1 were significantly upregulated in the livers of old mice relative to young mice and this increase occurred in the later half of life (i.e., after 18 months of age). Markers of M1 macrophages, expression of pro‐inflammatory cytokines (TNFα, IL6 and IL1β), and markers of fibrosis were all significantly upregulated in the liver with age and the change in necroptosis paralleled the changes in inflammation and fibrosis. Hepatocytes and liver macrophages isolated from old mice showed elevated levels of necroptosis markers as well as increased expression of pro‐inflammatory cytokines relative to young mice. Short‐term treatment with the necroptosis inhibitor, necrostatin‐1s (Nec‐1s), reduced necroptosis, markers of M1 macrophages, fibrosis, and cell senescence as well as reducing the expression of pro‐inflammatory cytokines in the livers of old mice. Thus, our data show for the first time that liver aging is associated with increased necroptosis and necroptosis contributes to chronic inflammation in the liver, which in turn appears to contribute to liver fibrosis and possibly CLD.     

Increased in vivo mitochondrial oxygenation with right ventricular failure induced by pulmonary arterial hypertension: mitochondrial inhibition as driver of cardiac failure?

Background

The leading cause of mortality due to pulmonary arterial hypertension (PAH) is failure of the cardiac right ventricle. It has long been hypothesized that during the development of chronic cardiac failure the heart becomes energy deprived, possibly due to shortage of oxygen at the level of cardiomyocyte mitochondria. However, direct evaluation of oxygen tension levels within the in vivo right ventricle during PAH is currently lacking. Here we directly evaluated this hypothesis by using a recently reported technique of oxygen-dependent quenching of delayed fluorescence of mitochondrial protoprophyrin IX, to determine the distribution of mitochondrial oxygen tension (mitoPO₂) within the right ventricle (RV) subjected to progressive PAH.

Methods

PAH was induced through a single injection of monocrotaline (MCT). Control (saline-injected), compensated RV hypertrophy (30 mg/kg MCT; MCT30), and RV failure (60 mg/kg MCT; MCT60) rats were compared 4 wk after treatment. The distribution of mitoPO₂ within the RV was determined in mechanically-ventilated, anaesthetized animals, applying different inspired oxygen (FiO₂) levels and two increment dosages of dobutamine.

Results

MCT60 resulted in RV failure (increased mortality, weight loss, increased lung weight), MCT30 resulted in compensated RV hypertrophy. At 30% or 40% FiO₂, necessary to obtain physiological arterial PO₂ in the diseased animals, RV failure rats had significantly less mitochondria (15% of total mitochondria) in the 0-20 mmHg mitoPO₂ range than hypertrophied RV rats (48%) or control rats (54%). Only when oxygen supply was reduced to 21% FiO_2, resulting in low arterial PO₂ for the MCT60 animals, or when oxygen demand increased with high dose dobutamine, the number of failing RV mitochondria with low oxygen became similar to control RV. In addition, metabolic enzyme analysis revealed similar mitochondrial mass, increased glycolytic hexokinase activity following MCT, with increased lactate dehydrogenase activity only in compensated hypertrophied RV.

Conclusions

Our novel observation of increased mitochondrial oxygenation suggests down-regulation of in vivo mitochondrial oxygen consumption, in the absence of hypoxia, with transition towards right ventricular failure induced by pulmonary arterial hypertension.     

Phosphoproteomic screening identifies Rab GTPases as novel downstream targets of PINK1

Mutations in the PTEN‐induced kinase 1 (PINK1) are causative of autosomal recessive Parkinson''s disease (PD). We have previously reported that PINK1 is activated by mitochondrial depolarisation and phosphorylates serine 65 (Ser⁶⁵) of the ubiquitin ligase Parkin and ubiquitin to stimulate Parkin E3 ligase activity. Here, we have employed quantitative phosphoproteomics to search for novel PINK1‐dependent phosphorylation targets in HEK (human embryonic kidney) 293 cells stimulated by mitochondrial depolarisation. This led to the identification of 14,213 phosphosites from 4,499 gene products. Whilst most phosphosites were unaffected, we strikingly observed three members of a sub‐family of Rab GTPases namely Rab8A, 8B and 13 that are all phosphorylated at the highly conserved residue of serine 111 (Ser¹¹¹) in response to PINK1 activation. Using phospho‐specific antibodies raised against Ser¹¹¹ of each of the Rabs, we demonstrate that Rab Ser¹¹¹ phosphorylation occurs specifically in response to PINK1 activation and is abolished in HeLa PINK1 knockout cells and mutant PINK1 PD patient‐derived fibroblasts stimulated by mitochondrial depolarisation. We provide evidence that Rab8A GTPase Ser¹¹¹ phosphorylation is not directly regulated by PINK1 in vitro and demonstrate in cells the time course of Ser¹¹¹ phosphorylation of Rab8A, 8B and 13 is markedly delayed compared to phosphorylation of Parkin at Ser⁶⁵. We further show mechanistically that phosphorylation at Ser¹¹¹ significantly impairs Rab8A activation by its cognate guanine nucleotide exchange factor (GEF), Rabin8 (by using the Ser111Glu phosphorylation mimic). These findings provide the first evidence that PINK1 is able to regulate the phosphorylation of Rab GTPases and indicate that monitoring phosphorylation of Rab8A/8B/13 at Ser¹¹¹ may represent novel biomarkers of PINK1 activity in vivo. Our findings also suggest that disruption of Rab GTPase‐mediated signalling may represent a major mechanism in the neurodegenerative cascade of Parkinson''s disease.     

Dynamics of NK,CD8 and Tfh cell mediated the production of cytokines and antiviral antibodies in Chinese patients with moderate COVID‐19

Recent studies have demonstrated a marked decrease in peripheral lymphocyte levels in patients with coronavirus disease 2019 (COVID‐19) caused by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2). Few studies have focused on the changes of NK, T‐ and B‐cell subsets, inflammatory cytokines and virus‐specific antibodies in patients with moderate COVID‐19. A total of 11 RT‐PCR‐confirmed convalescent patients with COVID‐19 and 11 patients with non‐SARS‐CoV‐2 pneumonia (control patients) were enrolled in this study. NK, CD8⁺ T, CD4⁺ T, Tfh‐like and B‐cell subsets were analysed using flow cytometry. Cytokines and SARS‐CoV‐2‐specific antibodies were analysed using an electrochemiluminescence immunoassay. NK cell counts were significantly higher in patients with COVID‐19 than in control patients (P = 0.017). Effector memory CD8⁺ T‐cell counts significantly increased in patients with COVID‐19 during a convalescent period of 1 week (P = 0.041). TIM‐3⁺ Tfh‐like cell and CD226⁺ Tfh‐like cell counts significantly increased (P = 0.027) and decreased (P = 0.022), respectively, during the same period. Moreover, ICOS⁺ Tfh‐like cell counts tended to decrease (P = 0.074). No abnormal increase in cytokine levels was observed. The high expression of NK cells is important in innate immune response against SARS‐CoV‐2. The increase in effector memory CD8⁺ T‐cell counts, the up‐regulation of inhibitory molecules and the down‐regulation of active molecules on CD4⁺ T cells and Tfh‐like cells in patients with COVID‐19 would benefit the maintenance of balanced cellular and humoural immune responses, may prevent the development of severe cases and contribute to the recovery of patients with COVID‐19.     

SARS‐CoV‐2 nucleocapsid suppresses host pyroptosis by blocking Gasdermin D cleavage

SARS‐CoV‐2 is an emerging coronavirus that causes dysfunctions in multiple human cells and tissues. Studies have looked at the entry of SARS‐CoV‐2 into host cells mediated by the viral spike protein and human receptor ACE2. However, less is known about the cellular immune responses triggered by SARS‐CoV‐2 viral proteins. Here, we show that the nucleocapsid of SARS‐CoV‐2 inhibits host pyroptosis by blocking Gasdermin D (GSDMD) cleavage. SARS‐CoV‐2‐infected monocytes show enhanced cellular interleukin‐1β (IL‐1β) expression, but reduced IL‐1β secretion. While SARS‐CoV‐2 infection promotes activation of the NLRP3 inflammasome and caspase‐1, GSDMD cleavage and pyroptosis are inhibited in infected human monocytes. SARS‐CoV‐2 nucleocapsid protein associates with GSDMD in cells and inhibits GSDMD cleavage in vitro and in vivo. The nucleocapsid binds the GSDMD linker region and hinders GSDMD processing by caspase‐1. These insights into how SARS‐CoV‐2 antagonizes cellular inflammatory responses may open new avenues for treating COVID‐19 in the future.     

IFN‐γ+IL‐17+Th17 cells regulate fibrosis through secreting IL‐21 in systemic scleroderma

This study aimed to explore the function of IFN‐γ⁺IL‐17⁺Th17 cells on fibrosis in systemic scleroderma (SSc). Blood and skin samples were collected from 20 SSc cases and 10 healthy individuals. The percentage of IFN‐γ⁺IL‐17⁺Th17 cells was detected using flow cytometry. The in vitro induction of IFN‐γ⁺IL‐17⁺Th17 cells was performed adopting PHA and rIL‐12. Gene expression was detected via quantitative real‐time polymerase chain reaction (qRT‐PCR), whereas western blot analysis was adopted for protein analysis. The distribution of IFN‐γ⁺IL‐17⁺Th17 cells was significantly increased in SSc cases and positively correlated with SSc stages (P = .031), disease duration (P = .016), activity (P = .025) and skin scores (P < .001). In vitro, IFN‐γ⁺IL‐17⁺Th17 cells could promote the expressions of α‐SMA and COL1A1, revealing increased fibroblasts’ proliferation and enhanced collagen‐secreting capacity. In addition, IL‐21 expression was significantly increased in co‐culture medium of IFN‐γ⁺IL‐17⁺Th17 cells and fibroblasts (P < .001). IL‐21 neutralizer treatment resulted in the down‐regulation of α‐SMA and COL1A1. IL‐21 was confirmed as an effector of IFN‐γ⁺IL‐17⁺Th17 cells in fibrosis process. The distribution of IFN‐γ⁺IL‐17⁺Th17 cells was significantly increased in SSc cases and positively correlated with disease activity. IFN‐γ⁺IL‐17⁺Th17 cells could promote fibroblast proliferation and enhance collagen‐secreting ability via producing IL‐21, thus contributing to fibrosis in SSc.     

Safety and feasibility of umbilical cord mesenchymal stem cells in patients with COVID‐19 pneumonia: A pilot study

ObjectivesWe aim to explore the safety and feasibility of umbilical cord mesenchymal stem cells (UC‐MSCs) transplantation in patients with severe and critically severe coronavirus disease‐2019 (COVID‐19).MethodsWe conducted a small sample, single arm, pilot trial. In addition to standard therapy, we performed four rounds of transplantation of UC‐MSCs in sixteen patients with severe and critically severe COVID‐19. We recorded adverse events from enrolment to Day 28. We evaluated the oxygenation index, inflammatory biomarkers, radiological presentations of the disease and lymphocyte subsets count on the 7th day (D7 ± 1 day), the 14th day (D14 ± 1 day) and the 28th day (D28 ± 3 days).ResultsThere were no infusion‐related or allergic reactions. The oxygenation index was improved after transplantation. The mortality of enrolled patients was 6.25%, whereas the historical mortality rate was 45.4%. The level of cytokines estimated varied in the normal range, the radiological presentations (ground glass opacity) were improved and the lymphocyte count and lymphocyte subsets (CD4⁺ T cells, CD8⁺ T cells and NK cells) count showed recovery after transplantation.ConclusionsIntravenous transplantation of UC‐MSCs was safe and feasible for treatment of patients with severe and critically severe COVID‐19 pneumonia.     

Genotyping validates the efficacy of photographic identification in a capture‐mark‐recapture study based on the head scale patterns of the prairie lizard (Sceloporus consobrinus)

Population studies often incorporate capture‐mark‐recapture (CMR) techniques to gather information on long‐term biological and demographic characteristics. A fundamental requirement for CMR studies is that an individual must be uniquely and permanently marked to ensure reliable reidentification throughout its lifespan. Photographic identification involving automated photographic identification software has become a popular and efficient noninvasive method for identifying individuals based on natural markings. However, few studies have (a) robustly assessed the performance of automated programs by using a double‐marking system or (b) determined their efficacy for long‐term studies by incorporating multi‐year data. Here, we evaluated the performance of the program Interactive Individual Identification System (I³S) by cross‐validating photographic identifications based on the head scale pattern of the prairie lizard (Sceloporus consobrinus) with individual microsatellite genotyping (N = 863). Further, we assessed the efficacy of the program to identify individuals over time by comparing error rates between within‐year and between‐year recaptures. Recaptured lizards were correctly identified by I³S in 94.1% of cases. We estimated a false rejection rate (FRR) of 5.9% and a false acceptance rate (FAR) of 0%. By using I³S, we correctly identified 97.8% of within‐year recaptures (FRR = 2.2%; FAR = 0%) and 91.1% of between‐year recaptures (FRR = 8.9%; FAR = 0%). Misidentifications were primarily due to poor photograph quality (N = 4). However, two misidentifications were caused by indistinct scale configuration due to scale damage (N = 1) and ontogenetic changes in head scalation between capture events (N = 1). We conclude that automated photographic identification based on head scale patterns is a reliable and accurate method for identifying individuals over time. Because many lizard or reptilian species possess variable head squamation, this method has potential for successful application in many species.     

Dexamethasone induces apoptosis in pulmonary arterial smooth muscle cells

Background

Dexamethasone suppressed inflammation and haemodynamic changes in an animal model of pulmonary arterial hypertension (PAH). A major target for dexamethasone actions is NF-κB, which is activated in pulmonary vascular cells and perivascular inflammatory cells in PAH. Reverse remodelling is an important concept in PAH disease therapy, and further to its anti-proliferative effects, we sought to explore whether dexamethasone augments pulmonary arterial smooth muscle cell (PASMC) apoptosis.

Methods

Analysis of apoptosis markers (caspase 3, in-situ DNA fragmentation) and NF-κB (p65 and phospho-IKK-α/β) activation was performed on lung tissue from rats with monocrotaline (MCT)-induced pulmonary hypertension (PH), before and after day 14–28 treatment with dexamethasone (5 mg/kg/day). PASMC were cultured from this rat PH model and from normal human lung following lung cancer surgery. Following stimulation with TNF-α (10 ng/ml), the effects of dexamethasone (10⁻⁸–10⁻⁶ M) and IKK2 (NF-κB) inhibition (AS602868, 0–3 μM (0-3×10⁻⁶ M) on IL-6 and CXCL8 release and apoptosis was determined by ELISA and by Hoechst staining. NF-κB activation was measured by TransAm assay.

Results

Dexamethasone treatment of rats with MCT-induced PH in vivo led to PASMC apoptosis as displayed by increased caspase 3 expression and DNA fragmentation. A similar effect was seen in vitro using TNF-α-simulated human and rat PASMC following both dexamethasone and IKK2 inhibition. Increased apoptosis was associated with a reduction in NF-κB activation and in IL-6 and CXCL8 release from PASMC.

Conclusions

Dexamethasone exerted reverse-remodelling effects by augmenting apoptosis and reversing inflammation in PASMC possibly via inhibition of NF-κB. Future PAH therapies may involve targeting these important inflammatory pathways.     

Regulatory KIR+RA+ T cells accumulate with age and are highly activated during viral respiratory disease

Severe respiratory viral infectious diseases such as influenza and COVID‐19 especially affect the older population. This is partly ascribed to diminished CD8⁺ T‐cell responses a result of aging. The phenotypical diversity of the CD8⁺ T‐cell population has made it difficult to identify the impact of aging on CD8⁺ T‐cell subsets associated with diminished CD8⁺ T‐cell responses. Here we identify a novel human CD8⁺ T‐cell subset characterized by expression of Killer‐cell Immunoglobulin‐like Receptors (KIR⁺) and CD45RA (RA⁺). These KIR⁺RA⁺ T cells accumulated with age in the blood of healthy individuals (20–82 years of age, n = 50), expressed high levels of aging‐related markers of T‐cell regulation, and were functionally capable of suppressing proliferation of other CD8⁺ T cells. Moreover, KIR⁺RA⁺ T cells were a major T‐cell subset becoming activated in older adults suffering from an acute respiratory viral infection (n = 36), including coronavirus and influenza virus infection. In addition, older adults with influenza A infection showed that higher activation status of their KIR⁺RA⁺ T cells associated with longer duration of respiratory symptoms. Together, our data indicate that KIR⁺RA⁺ T cells are a unique human T‐cell subset with regulatory properties that may explain susceptibility to viral respiratory disease at old age.     

Camera trap reveals the co‐occurrence patterns of two sympatric muntjac species in southern Anhui Province,China: No spatial segregation

The competitive relationship and coexistence pattern among close related species have long been one of the hot issues in ecological research. Interspecies interactions can exert important influences on the local distribution of rare species. Black muntjac Muntiacus crinifrons is an endemic species to eastern China, currently restricted to limited regions. In contrast, Chinese muntjac Muntiacus reevesi is the most common and widespread deer in southern China. Both species co‐occur in southern Anhui and western Zhejiang Province. Little is known about the interaction of these two sympatric‐related species. In this study, to investigate the site use determinants and co‐occurrence pattern of the two sympatric muntjac species, we conducted a camera trap survey across about 250 km² in mountainous area of southern Anhui Province, China. We adopted a multistep approach to incorporate habitat preferences while modeling occupancy and detection. We found that the two species did not separate along elevation gradient (range from 400 m to 1,400 m) as described in previous studies. Results of single‐species occupancy models indicated that elevation had positive effects on the site use of both species, while slope had an opposite influence on their site use. Positive effects of elevation on the site use implied that both species try to avoid human interference at low elevations. Significant negative effect of slope on the site use of black muntjac suggested that the species prefer habitat with gentle slope and avoided steep. Co‐occurrence models and species interaction factors provided evidence that the two muntjac species had an independent occupancy (ψ ^{BM CM} = ψ ^{BM cm}, SIF = 1) and exhibited a positive species interaction in detection probability (p ^BM < r ^{BM CM}). Combined with the results of previous studies, we suggested that it was fine differentiation in microhabitats and food resources utilization rather spatial or temporal segregation that allowed the two species co‐occurrence. The site use determinants revealed in our study would be useful for the habitat conservation and restoration for the rare black muntjac, and the co‐occurrence pattern of the two sympatric muntjac species could provide useful information for deep understanding of the coexistence mechanism among forest‐dwelling ungulates.     

ERK/Drp1‐dependent mitochondrial fission contributes to HMGB1‐induced autophagy in pulmonary arterial hypertension

ObjectivesHigh‐mobility group box‐1 (HMGB1) and aberrant mitochondrial fission mediated by excessive activation of GTPase dynamin‐related protein 1 (Drp1) have been found to be elevated in patients with pulmonary arterial hypertension (PAH) and critically implicated in PAH pathogenesis. However, it remains unknown whether Drp1‐mediated mitochondrial fission and which downstream targets of mitochondrial fission mediate HMGB1‐induced pulmonary arterial smooth muscle cells (PASMCs) proliferation and migration leading to vascular remodelling in PAH. This study aims to address these issues.MethodsPrimary cultured PASMCs were obtained from male Sprague‐Dawley (SD) rats. We detected RNA levels by qRT‐PCR, protein levels by Western blotting, cell proliferation by Cell Counting Kit‐8 (CCK‐8) and EdU incorporation assays, migration by wound healing and transwell assays. SD rats were injected with monocrotaline (MCT) to establish PAH. Hemodynamic parameters were measured by closed‐chest right heart catheterization.ResultsHMGB1 increased Drp1 phosphorylation and Drp1‐dependent mitochondrial fragmentation through extracellular signal‐regulated kinases 1/2 (ERK1/2) signalling activation, and subsequently triggered autophagy activation, which further led to bone morphogenetic protein receptor 2 (BMPR2) lysosomal degradation and inhibitor of DNA binding 1 (Id1) downregulation, and eventually promoted PASMCs proliferation/migration. Inhibition of ERK1/2 cascade, knockdown of Drp1 or suppression of autophagy restored HMGB1‐induced reductions of BMPR2 and Id1, and diminished HMGB1‐induced PASMCs proliferation/migration. In addition, pharmacological inhibition of HMGB1 by glycyrrhizin, suppression of mitochondrial fission by Mdivi‐1 or blockage of autophagy by chloroquine prevented PAH development in MCT‐induced rats PAH model.ConclusionsHMGB1 promotes PASMCs proliferation/migration and pulmonary vascular remodelling by activating ERK1/2/Drp1/Autophagy/BMPR2/Id1 axis, suggesting that this cascade might be a potential novel target for management of PAH.     

Characterization of the glutathione‐dependent reduction of the peroxiredoxin 5 homolog PfAOP from Plasmodium falciparum

Peroxiredoxins use a variety of thiols to rapidly reduce hydroperoxides and peroxynitrite. While the oxidation kinetics of peroxiredoxins have been studied in great detail, enzyme‐specific differences regarding peroxiredoxin reduction and the overall rate‐limiting step under physiological conditions often remain to be deciphered. The 1‐Cys peroxiredoxin 5 homolog PfAOP from the malaria parasite Plasmodium falciparum is an established model enzyme for glutathione/glutaredoxin‐dependent peroxiredoxins. Here, we reconstituted the catalytic cycle of PfAOP in vitro and analyzed the reaction between oxidized PfAOP and reduced glutathione (GSH) using molecular docking and stopped‐flow measurements. Molecular docking revealed that oxidized PfAOP has to adopt a locally unfolded conformation to react with GSH. Furthermore, we determined a second‐order rate constant of 6 × 10⁵ M⁻¹ s⁻¹ at 25°C and thermodynamic activation parameters ΔH ^‡, ΔS ^‡, and ΔG ^‡ of 39.8 kJ/mol, −0.8 J/mol, and 40.0 kJ/mol, respectively. The gain‐of‐function mutant PfAOP^L109M had almost identical reaction parameters. Taking into account physiological hydroperoxide and GSH concentrations, we suggest (a) that the reaction between oxidized PfAOP and GSH might be even faster than the formation of the sulfenic acid in vivo, and (b) that conformational changes are likely rate limiting for PfAOP catalysis. In summary, we characterized and quantified the reaction between GSH and the model enzyme PfAOP, thus providing detailed insights regarding the reactivity of its sulfenic acid and the versatile chemistry of peroxiredoxins.     

Optimal foraging by a large ungulate in an extreme environment: Wild mountain reindeer select snow‐free feeding habitats in winter

Optimal foraging models predict that individual animals will optimize net energy gain by intensifying forage activity and/or reducing forage energy cost. Then, the free distribution model predicts an animal''s distribution in a patchy landscape will match the distribution of the resources. If not modified by other factors, such patterns may be expected to be particularly explicit in variable and extreme, forage‐limited, and patchy environments, notably alpine and Arctic environments during winter. The large ungulate wild mountain reindeer (Rangifer tarandus tarandus) surviving in such environments is used as a model during the forage‐limited winter season. The largest wild reindeer area in Western Europe (Hardangervidda, 8130 km²) is actively managed to sustain 10,000–12,000 wild reindeer. Since 2001, 104 different individuals have been GPS‐tracked at 3‐hr intervals. In winter, mountain reindeer may either choose to seek out and forage in patchy snow‐free habitats, typically on top of wind‐blown ridges, or use energy‐demanding digging through the snow to reach ground forage (cratering). We use late April satellite data from Landsat 5 and 8 (30 × 30 m), airborne laser scanning subsampling (processed to 1 × 1 m grid), and topographic information (1 m resolution) derived from digital aerial photographs (0.25 × 0.25 m resolution) to delineate snow‐free patches, constituting less than 694 km². By overlaying recorded wild reindeer GPS positions winters 2001–2017 (188,942 positions), we document a strong positive selection for snow‐free patches, which were used about four times more frequently than expected from a “random walk” model. On a daily basis, the preference for snow‐free areas was slightly stronger in the evenings. In the sustainable management of wild mountain reindeer, the area of snow‐free patches is an important predictor of winter forage availability and important winter source areas. It may be derived from remote sensing data.     

Allogeneic adipose tissue‐derived stem cells ELIXCYTE® in chronic kidney disease: A phase I study assessing safety and clinical feasibility

The purpose of this phase I clinical trial is to assess the safety and tolerability of allogeneic adipose tissue‐derived stem cells (ADSCs) among chronic kidney disease (CKD) patients. 12 eligible CKD patients with an estimated glomerular filtration rate (eGFR) of 15–44 ml/min/1.73 m² received one dose of intravenous allogeneic ADSCs (ELIXCYTE^®), as 3 groups: 3 low dose (6.4 × 10⁷ cells in total of 8 ml), 3 middle dose (19.2 × 10⁷ cells in total of 24 ml) and 6 high dose (32.0 × 10⁷ cells in total of 40 ml) of ELIXCYTE^® and evaluated after 48 weeks. Primary endpoint was the safety profiles in terms of incidence of adverse events (AEs) and serious adverse event (SAE). Two subjects in high dose group experienced a total of 2 treatment‐related AEs which are Grade 1 slow speech and Grade 1 bradyphrenia after the infusion. One subject in middle dose group experienced an SAE unlikely related to treatment, grade 2 proteinuria. No fatal AE was reported in this study. An increase in eGFR was observed in 7 out of 12 subjects (58%) at Week 24 and in 6 of 12 subjects (50%) by Week 48. By Week 24, an increase in eGFR by more than 20% among all CKD patients with baseline eGFR ≧ 30 ml/min/1.73 m² as compared to only 2 subjects in baseline eGFR < 30 ml/min/1.73 m² group. No significant reduction in proteinuria was noted among all subjects. This phase I trial demonstrated single‐dose intravenous ELIXCYTE was well tolerated in moderate‐to‐severe CKD patients and its preliminary efficacy warrants future studies.     

Genetic trade‐offs between complex diseases and longevity

Longevity was influenced by many complex diseases and traits. However, the relationships between human longevity and genetic risks of complex diseases were not broadly studied. Here, we constructed polygenic risk scores (PRSs) for 225 complex diseases/traits and evaluated their relationships with human longevity in a cohort with 2178 centenarians and 2299 middle‐aged individuals. Lower genetic risks of stroke and hypotension were observed in centenarians, while higher genetic risks of schizophrenia (SCZ) and type 2 diabetes (T2D) were detected in long‐lived individuals. We further stratified PRSs into cell‐type groups and significance‐level groups. The results showed that the immune component of SCZ genetic risk was positively linked to longevity, and the renal component of T2D genetic risk was the most deleterious. Additionally, SNPs with very small p‐values (p ≤ 1x10^‐5) for SCZ and T2D were negatively correlated with longevity. While for the less significant SNPs (1x10^‐5 < p ≤ 0.05), their effects on disease and longevity were positively correlated. Overall, we identified genetically informed positive and negative factors for human longevity, gained more insights on the accumulation of disease risk alleles during evolution, and provided evidence for the theory of genetic trade‐offs between complex diseases and longevity.