Absence of PD-L1 signaling hinders macrophage defense against Mycobacterium tuberculosis via upregulating STAT3/IL-6 pathway.

The blockade of programmed death-ligand 1 (PD-L1) pathway has been clinically used in cancer immunotherapy, while its effects on infectious diseases remain elusive. Roles of PD-L1 signaling in the macrophage-mediated innate immune defense against M.tb is unclear. In this study, the outcomes of tuberculosis (TB) in wild-type (WT) mice treated with anti-PD-1/PD-L1 therapy and macrophage-specific Pdl1-knockout (Pdl1ΔΜΦ) mice were compared. Treatment with anti-PD-L1 or anti-PD-1 benefited protection against M.tb infection in WT mice, while Pdl1ΔΜΦ mice exhibited the increased susceptibility to M.tb infection. Mechanistically, the absence of PD-L1 signaling impaired M.tb killing by macrophages. Furthermore, elevated STAT3 activation was found in PD-L1-deficient macrophages, leading to increased interleukin (IL)-6 production and reduced inducible nitric oxide synthase (iNOS) expression. Inhibiting STAT3 phosphorylation partially impeded the increase in IL-6 production and restored iNOS expression in these PD-L1-deficient cells. These findings provide valuable insights into the complexity and mechanisms underlying anti-PD-L1 therapy in the context of tuberculosis.

Intrauterine Growth Restriction Affects Colonic Barrier Function via Regulating the Nrf2/Keap1 and TLR4-NF-κB/ERK Pathways and Altering Colonic Microbiome and Metabolome Homeostasis in Growing-Finishing Pigs.

Intrauterine growth restriction (IUGR) pigs are characterized by long-term growth failure, metabolic disorders, and intestinal microbiota imbalance. The characteristics of the negative effects of IUGR at different growth stages of pigs are still unclear. Therefore, this study explored through multi-omics analyses whether the IUGR damages the intestinal barrier function and alters the colonization and metabolic profiles of the colonic microbiota in growing-finishing pigs. Seventy-two piglets (36 IUGR and 36 NBW) were allocated for this trial to analyze physiological and plasma biochemical parameters, as well as oxidative damage and inflammatory response in the colon. Moreover, the colonic microbiota communities and metabolome were examined using 16s rRNA sequencing and metabolomics technologies to reveal the intestinal characteristics of IUGR pigs at different growth stages (25, 50, and 100 kg). IUGR altered the concentrations of plasma glucose, total protein, triglycerides, and cholesterol. Colonic tight junction proteins were markedly inhibited by IUGR. IUGR decreased plasma T-AOC, SOD, and GSH levels and colonic SOD-1, SOD-2, and GPX-4 expressions by restraining the Nrf2/Keap1 signaling pathway. Moreover, IUGR increased colonic IL-1ß and TNF-α levels while reducing IL-10, possibly through activating the TLR4-NF-κB/ERK pathway. Notably, IUGR pigs had lower colonic Streptococcus abundance and Firmicutes-to-Bacteroidetes ratio at the 25 kg BW stage while having higher Firmicutes abundance at the 100 kg BW stage; moreover, IUGR pigs had lower SCFA concentrations. Metabolomics analysis showed that IUGR increased colonic lipids and lipid-like molecules, organic acids and derivatives, and organoheterocyclic compounds concentrations and enriched three differential metabolic pathways, including linoleic acid, sphingolipid, and purine metabolisms throughout the trial. Collectively, IUGR altered the nutrient metabolism, redox status, and colonic microbiota community and metabolite profiles of pigs and continued to disrupt colonic barrier function by reducing antioxidant capacity via the Nrf2/Keap1 pathway and activating inflammation via the TLR4-NF-κB/ERK pathway during the growing-finishing stage. Moreover, colonic Firmicutes and Streptococcus could be potential regulatory targets for modulating the metabolism and health of IUGR pigs.

Maternal Supplementation with Ornithine Promotes Placental Angiogenesis and Improves Intestinal Development of Suckling Piglets.

The blood vessels of the placenta are crucial for fetal growth. Here, lower vessel density and ornithine (Orn) content were observed in placentae for low-birth-weight fetuses versus normal-birth-weight fetuses at day 75 of gestation. Furthermore, the Orn content in placentae decreased from day 75 to 110 of gestation. To investigate the role of Orn in placental angiogenesis, 48 gilts (Bama pig) were allocated into four groups. The gilts in the control group were fed a basal diet (CON group), while those in the experimental groups were fed a basal diet supplemented with 0.05% Orn (0.05% Orn group), 0.10% Orn (0.10% Orn group), and 0.15% Orn (0.15% Orn group), respectively. The results showed that 0.15% Orn and 0.10% Orn groups exhibited increased birth weight of piglets compared with the CON group. Moreover, the 0.15% Orn group was higher than the CON group in the blood vessel densities of placenta. Mechanistically, Orn facilitated placental angiogenesis by regulating vascular endothelial growth factor-A (VEGF-A). Furthermore, maternal supplementation with 0.15% Orn during gestation increased the jejunal and ileal villi height and the concentrations of colonic propionate and butyrate in suckling piglets. Collectively, these results showed that maternal supplementation with Orn promotes placental angiogenesis and improves intestinal development of suckling piglets.

Pericytes contribute to pulmonary vascular remodeling via HIF2α signaling.

Vascular remodeling is the process of structural alteration and cell rearrangement of blood vessels in response to injury and is the cause of many of the world's most afflicted cardiovascular conditions, including pulmonary arterial hypertension (PAH). Many studies have focused on the effects of vascular endothelial cells and smooth muscle cells (SMCs) during vascular remodeling, but pericytes, an indispensable cell population residing largely in capillaries, are ignored in this maladaptive process. Here, we report that hypoxia-inducible factor 2α (HIF2α) expression is increased in the lung tissues of PAH patients, and HIF2α overexpressed pericytes result in greater contractility and an impaired endothelial-pericyte interaction. Using single-cell RNAseq and hypoxia-induced pulmonary hypertension (PH) models, we show that HIF2α is a major molecular regulator for the transformation of pericytes into SMC-like cells. Pericyte-selective HIF2α overexpression in mice exacerbates PH and right ventricular hypertrophy. Temporal cellular lineage tracing shows that HIF2α overexpressing reporter NG2+ cells (pericyte-selective) relocate from capillaries to arterioles and co-express SMA. This novel insight into the crucial role of NG2+ pericytes in pulmonary vascular remodeling via HIF2α signaling suggests a potential drug target for PH.

MiR-488-3p facilitates wound healing through CYP1B1-mediated Wnt/ß-catenin signaling pathway by targeting MeCP2.

INTRODUCTION: Diabetic wounds are difficult to heal, but the pathogenesis is unknown. MicroRNAs (miRNAs) are thought to play important roles in wound healing. The effect of miR-488-3p in wound healing was studied in this article. MATERIALS AND METHODS: The gene methylation was measured by methylation specific PCR (MSP) assay. A dual-luciferase reporter assay was adopted to analyze the interaction between miR-488-3p and MeCP2. RESULTS: Cytochrome P450 1B1 (CYP1B1) is a monooxygenase belonging to the cytochrome P450 family that aids in wound healing. Our findings showed that the miR-488-3p and CYP1B1 expression levels were much lower in wound tissues of diabetics with skin defects, but the methyl-CpG-binding protein 2 (MeCP2) level was significantly higher than that in control skin tissues. MiR-488-3p overexpression increased cell proliferation and migration, as well as HUVEC angiogenesis, while inhibiting apoptosis, according to function experiments. In vitro, MeCP2 inhibited wound healing by acting as a target of miR-488-3p. We later discovered that MeCP2 inhibited CYP1B1 expression by enhancing its methylation state. In addition, CYP1B1 knockdown inhibited wound healing. Furthermore, MeCP2 overexpression abolished the promoting effect of miR-488-3p on wound healing. It also turned out that CYP1B1 promoted wound healing by activating the Wnt4/ß-catenin pathway. Animal experiments also showed that miR-488-3p overexpression could accelerate wound healing in diabetic male SD rats. CONCLUSIONS: MiR-488-3p is a potential therapeutic target for diabetic wound healing since it improved wound healing by activating the CYP1B1-mediated Wnt4/-catenin signaling cascade via MeCP2.

Putrescine promotes MMP9-induced angiogenesis in skeletal muscle through hydrogen peroxide/METTL3 pathway.

Blood vessels play a crucial role in the development of skeletal muscle, ensuring the supply of nutrients and oxygen. Putrescine, an essential polyamine for eukaryotic cells, has an unclear impact on skeletal muscle angiogenesis. In this study, we observed lower vessel density and reduced putrescine level in the muscle of low-birth-weight piglet models, and identified a positive correlation between putrescine content and muscle vessel density. Furthermore, putrescine was found to promote angiogenesis in skeletal muscle both in vitro and in vivo by targeting matrix metalloproteinase 9 (MMP9). On a mechanistic level, putrescine augmented the expression of methyltransferase like 3 (METTL3) by attenuating hydrogen peroxide production, thereby increasing the level of N6-methyladenosine (m6A)-modified MMP9 mRNA. This m6A-modified MMP9 mRNA was subsequently recognized and bound by the YTH N6-methyladenosine RNA binding protein 1 (YTHDF1), enhancing the stability of MMP9 mRNA and its protein expression, consequently accelerating angiogenesis in skeletal muscle. In summary, our findings suggest that putrescine enhances MMP9-mediated angiogenesis in skeletal muscle via the hydrogen peroxide/METTL3 pathway.

Case Report: Identification of a novel LYN::LINC01900 transcript with promyelocytic phenotype and TP53 mutation in acute myeloid leukemia.

Acute myeloid leukemia (AML) is a malignant disease of myeloid hematopoietic stem/progenitor cells characterized by the abnormal proliferation of primitive and naive random cells in the bone marrow and peripheral blood. Acute promyelocytic leukemia (APL) is a type (AML-M3) of AML. Most patients with APL have the characteristic chromosomal translocation t(15; 17)(q22; q12), forming PML::RARA fusion. The occurrence and progression of AML are often accompanied by the emergence of gene fusions such as PML::RARA, CBFß::MYH11, and RUNX1::RUNX1T1, among others. Gene fusions are the main molecular biological abnormalities in acute leukemia, and all fusion genes act as crucial oncogenic factors in leukemia. Herein, we report the first case of LYN::LINC01900 fusion transcript in AML with a promyelocytic phenotype and TP53 mutation. Further studies should address whether new protein products may result from this fusion, as well as the biological function of these new products in disease occurrence and progression.

An Unusual Anatomical Variation of Accessory Left Gastric Artery Arising from Left Hepatic Artery / Una Variación Anatómica Inusual de la Arteria Gástrica Izquierda Accesoria que se Origina de la Arteria Hepática Izquierda

SUMMARY: The stomach receives a rich blood supply from five sets of arteries, all of which originate from the celiac trunk. During the dissection of a female cadaver that had been fixed with formalin, an atypical branching pattern was observed. An accessory left gastric artery was found to originate from the left hepatic artery and send small branches to the esophagus, cardia, and fundus of the stomach. However, there was no anastomosis between the lower accessory left gastric artery and the left gastric artery. This is a rare variant of the gastric artery that has not been previously described in detail. It is important to recognize this variation for safe and effective interventional diagnosis and treatment techniques if dealing with the liver or gastric arteries.

El estómago recibe un rico suministro de sangre de cinco conjuntos de arterias, todas las cuales se originan en el tronco celíaco. Durante la disección de un cadáver femenino que había sido fijado con formalina, se observó un patrón de ramificación atípico. Se encontró una arteria gástrica izquierda accesoria que se originaba en la arteria hepática izquierda y enviaba pequeñas ramas al esófago, el cardias y el fondo del estómago. Sin embargo, no hubo anastomosis entre la arteria gástrica izquierda accesoria inferior y la arteria gástrica izquierda. Se trata de una variante rara de la arteria gástrica que no se ha descrito previamente en detalles. Es importante reconocer esta variación para la aplicación de técnicas de diagnóstico y tratamiento intervencionistas seguras y efectivas a nivel del hígado o las arterias gástricas.

DUOX2-Induced Oxidative Stress Inhibits Intestinal Angiogenesis through MMP3 in a Low-Birth-Weight Piglet Model.

Intestinal vessels play a critical role in nutrient absorption, whereas the effect and mechanism of low birth weight (LBW) on its formation remain unclear. Here, twenty newborn piglets were assigned to the control (CON) group (1162 ± 98 g) and LBW group (724 ± 31 g) according to their birth weight. Results showed that the villus height and the activity of maltase in the jejunum were lower in the LBW group than in the CON group. LBW group exhibited a higher oxidative stress level and impaired mitochondrial function in the jejunum and was lower than the CON group in the intestinal vascular density. To investigate the role of oxidative stress in intestinal angiogenesis, H2O2 was employed to induce oxidative stress in porcine intestinal epithelial cells (IPEC-J2). The results showed that the conditioned media from IPEC-J2 with H2O2 treatment decreased the angiogenesis of porcine vascular endothelial cells (PVEC). Transcriptome analysis revealed that a higher expression level of dual oxidase 2 (DUOX2) was found in the intestine of LBW piglets. Knockdown of DUOX2 in IPEC-J2 increased the proliferation and decreased the oxidative stress level. In addition, conditioned media from IPEC-J2 with DUOX2-knockdown was demonstrated to promote the angiogenesis of PVEC. Mechanistically, the knockdown of DUOX2 decreased the reactive oxygen species (ROS) level, thus increasing the angiogenesis in a matrix metalloproteinase 3 (MMP3) dependent manner. Conclusively, our results indicated that DUOX2-induced oxidative stress inhibited intestinal angiogenesis through MMP3 in a LBW piglet model.

Microenvironmental regulation of T-cells in pulmonary hypertension.

Introduction: In pulmonary hypertension (PH), pulmonary arterial remodeling is often accompanied by perivascular inflammation. The inflammation is characterized by the accumulation of activated macrophages and lymphocytes within the adventitial stroma, which is comprised primarily of fibroblasts. The well-known ability of fibroblasts to secrete interleukins and chemokines has previously been implicated as contributing to this tissue-specific inflammation in PH vessels. We were interested if pulmonary fibroblasts from PH arteries contribute to microenvironmental changes that could activate and polarize T-cells in PH. Methods: We used single-cell RNA sequencing of intact bovine distal pulmonary arteries (dPAs) from PH and control animals and flow cytometry, mRNA expression analysis, and respirometry analysis of blood-derived bovine/human T-cells exposed to conditioned media obtained from pulmonary fibroblasts of PH/control animals and IPAH/control patients (CM-(h)PH Fibs vs CM-(h)CO Fibs). Results: Single-cell RNA sequencing of intact bovine dPAs from PH and control animals revealed a pro-inflammatory phenotype of CD4+ T-cells and simultaneous absence of regulatory T-cells (FoxP3+ Tregs). By exposing T-cells to CM-(h)PH Fibs we stimulated their proinflammatory differentiation documented by increased IFNγ and decreased IL4, IL10, and TGFß mRNA and protein expression. Interestingly, we demonstrated a reduction in the number of suppressive T-cell subsets, i.e., human/bovine Tregs and bovine γδ T-cells treated with CM-(h)PH-Fibs. We also noted inhibition of anti-inflammatory cytokine expression (IL10, TGFß, IL4). Pro-inflammatory polarization of bovine T-cells exposed to CM-PH Fibs correlated with metabolic shift to glycolysis and lactate production with increased prooxidant intracellular status as well as increased proliferation of T-cells. To determine whether metabolic reprogramming of PH-Fibs was directly contributing to the effects of PH-Fibs conditioned media on T-cell polarization, we treated PH-Fibs with the HDAC inhibitor SAHA, which was previously shown to normalize metabolic status and examined the effects of the conditioned media. We observed significant suppression of inflammatory polarization associated with decreased T-cell proliferation and recovery of mitochondrial energy metabolism. Conclusion: This study demonstrates how the pulmonary fibroblast-derived microenvironment can activate and differentiate T-cells to trigger local inflammation, which is part of the vascular wall remodeling process in PH.

SIRT3 Is a Critical Regulator of Mitochondrial Function of Fibroblasts in Pulmonary Hypertension.

Pulmonary hypertension (PH) is a heterogeneous and life-threatening cardiopulmonary disorder in which mitochondrial dysfunction is believed to drive pathogenesis, although the underlying mechanisms remain unclear. To determine if abnormal SIRT3 (sirtuin 3) activity is related to mitochondrial dysfunction in adventitial fibroblasts from patients with idiopathic pulmonary arterial hypertension (IPAH) and hypoxic PH calves (PH-Fibs) and whether SIRT3 could be a potential therapeutic target to improve mitochondrial function, SIRT3 concentrations in control fibroblasts, PH-Fibs, and lung tissues were determined using quantitative real-time PCR and western blot. SIRT3 deacetylase activity in cells and lung tissues was determined using western blot, immunohistochemistry staining, and immunoprecipitation. Glycolysis and mitochondrial function in fibroblasts were measured using respiratory analysis and fluorescence-lifetime imaging microscopy. The effects of restoring SIRT3 activity (by overexpression of SIRT3 with plasmid, activation SIRT3 with honokiol, and supplementation with the SIRT3 cofactor nicotinamide adenine dinucleotide [NAD+]) on mitochondrial protein acetylation, mitochondrial function, cell proliferation, and gene expression in PH-Fibs were also investigated. We found that SIRT3 concentrations were decreased in PH-Fibs and PH lung tissues, and its cofactor, NAD+, was also decreased in PH-Fibs. Increased acetylation in overall mitochondrial proteins and SIRT3-specific targets (MPC1 [mitochondrial pyruvate carrier 1] and MnSOD2 [mitochondrial superoxide dismutase]), as well as decreased MnSOD2 activity, was identified in PH-Fibs and PH lung tissues. Normalization of SIRT3 activity, by increasing its expression with plasmid or with honokiol and supplementation with its cofactor NAD+, reduced mitochondrial protein acetylation, improved mitochondrial function, inhibited proliferation, and induced apoptosis in PH-Fibs. Thus, our study demonstrated that restoration of SIRT3 activity in PH-Fibs can reduce mitochondrial protein acetylation and restore mitochondrial function and PH-Fib phenotype in PH.

Effects of Dietary Probiotics and Acidifiers on the Production Performance, Colostrum Components, Serum Antioxidant Activity and Hormone Levels, and Gene Expression in Mammary Tissue of Lactating Sows.

The aims of this study were to test the effects of dietary probiotics and acidifiers on the production performance, colostrum components, serum antioxidant activity and hormone levels, and gene expression in the mammary tissue of lactating sows. Four treatments were administered with six replicates to 24 lactating sows. The control group (GC) received a basal diet, while the experimental groups received a basal diet with 200 mL/d probiotics (GP), 0.5% acidifiers (GA), and 200 mL/d probiotics + 0.5% acidifiers (GM), respectively. Compared with the GC, (1) the average weight of the piglets on the 21st day of lactation in the GM was higher (p < 0.05); (2) the colostrum fat ratio increased significantly (p < 0.05); (3) the malondialdehyde levels in GP and GM were lower (p < 0.05) on the 11th day; (4) on the 1st, 11th, and 21st days, the prolactin in GP and GM increased (p < 0.05); (5) on the 21st day, the relative expression levels of the prolactin receptor and fatty acid synthase were increased (p < 0.05). In summary, the basal diet mixed with 200 mL/d probiotics + 0.5% acidifiers could improve the production performance, colostrum components, serum antioxidant activity, and hormone levels of lactating sows.

Dietary Supplementation with Putrescine Improves Growth Performance and Meat Quality of Wenchang Chickens.

This study was to investigate the effects of dietary supplementation with putrescine on the growth performance and meat quality of chickens. A total of 480 eighty-day-old female Wenchang chickens were randomly assigned into four groups, with 8 replications per group and 15 animals per replicate. The chickens in the control group (Con) were fed a basal diet; the 3 experimental groups were fed a basal diet with 0.01%, 0.03%, and 0.05% putrescine, respectively. The experiment lasted for 40 days. The results showed that dietary supplementation with 0.05% putrescine increased (p < 0.05) the final body weight and average daily weight gain, and decreased the ratio of feed intake to the body weight gain of Wenchang chickens. Dietary supplementation with putrescine decreased the concentrations of putrescine, spermidine, and spermine in serum (p < 0.05). The contents of methionine, phenylalanine, lysine, aspartic acid, tyrosine, total essential amino acids, and total amino acids in breast muscle were higher (p < 0.05) in 0.03% and 0.05% groups than those in Con group. However, the contents of undecanoic acid, lauric acid, tridecanoic acid, myristic acid, pentadecanoic acid, arachidic acid, docosanoic acid, tricosanic acid, lignoceric acid, erucic acid, cis-11,14,17-eicosatrienoate, linoleic acid, and total n-6 monounsaturated fatty acids in breast muscle were lower (p < 0.05) in 0.03% and 0.05% groups than those in Con group. In addition, putrescine supplementation decreased (p < 0.05) the ratio of n-6/n-3 polyunsaturated fatty acids in breast meat. Overall, dietary supplementation with 0.05% putrescine enhanced the growth performance and meat quality of Wenchang chickens.

Efficacy and Safety of Photodynamic Therapy for the Treatment of Actinic Keratoses: A Meta-Analysis Update of Randomized Controlled Trials.

BACKGROUND: Actinic keratoses (AKs) are common and some may evolve into squamous cell carcinoma. Photodynamic therapy (PDT), imiquimod, cryotherapy, and other methods have been reported to have good effects. However, which treatment is the most effective with the best cosmetic results and fewest complications is uncertain. OBJECTIVE: To evaluate which method has the best efficacy and cosmetic results with less adverse events and recurrence rate. MATERIALS AND METHODS: All relevant articles up to July 31, 2022 were searched from Cochrane, Embase, and PubMed databases. Extract and analyze the data of efficacy, cosmetic results, local reactions, and adverse effects. RESULTS: Twenty-nine articles with 3,,850 participants and 24,747 lesions were included. Quality of evidence was generally high. The efficacy of PDT was better in complete response (CR) (lesions CR; risk ratio (RR) 1.87; 95% confidence interval (CI) 1.55-1.87/patient CR; RR 3.07; 95% CI 2.07-4.56), overall preference, and cosmetic results. The time cumulative meta-analysis showed that the curative effect was gradually increasing before 2004, and then gradually stabilizing. Two groups showed no statistically significant differences in recurrence. CONCLUSION: Compared with other methods, PDT is significantly more effective for AK with excellent cosmetic results and reversible adverse effects.

Local Complement Contributes to Pathogenic Activation of Lung Endothelial Cells in SARS-CoV-2 Infection.

Endothelial dysfunction and inflammation contribute to the vascular pathology of coronavirus disease (COVID-19). However, emerging evidence does not support direct infection of endothelial or other vascular wall cells, and thus inflammation may be better explained as a secondary response to epithelial cell infection. In this study, we sought to determine whether lung endothelial or other resident vascular cells are susceptible to productive severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and how local complement activation contributes to endothelial dysfunction and inflammation in response to hypoxia and SARS-CoV-2-infected lung alveolar epithelial cells. We found that ACE2 (angiotensin-converting enzyme 2) and TMPRSS2 (transmembrane serine protease 2) mRNA expression in lung vascular cells, including primary human lung microvascular endothelial cells (HLMVECs), pericytes, smooth muscle cells, and fibroblasts, was 20- to 90-fold lower compared with primary human alveolar epithelial type II cells. Consistently, we found that HLMVECs and other resident vascular cells were not susceptible to productive SARS-CoV-2 infection under either normoxic or hypoxic conditions. However, viral uptake without replication (abortive infection) was observed in HLMVECs when exposed to conditioned medium from SARS-CoV-2-infected human ACE2 stably transfected A549 epithelial cells. Furthermore, we demonstrated that exposure of HLMVECs to conditioned medium from SARS-CoV-2-infected human ACE2 stably transfected A549 epithelial cells and hypoxia resulted in upregulation of inflammatory factors such as ICAM-1 (intercellular adhesion molecule 1), VCAM-1 (vascular cell adhesion molecule 1), and IL-6 (interleukin 6) as well as complement components such as C3 (complement C3), C3AR1 (complement C3a receptor 1), C1QA (complement C1q A chain), and CFB (complement factor B). Taken together, our data support a model in which lung endothelial and vascular dysfunction during COVID-19 involves the activation of complement and inflammatory signaling and does not involve productive viral infection of endothelial cells.

Functional and molecular determinants of right ventricular response to severe pulmonary hypertension in a large animal model.

Right ventricular (RV) failure is the major determinant of outcome in pulmonary hypertension (PH). Calves exposed to 2-wk hypoxia develop severe PH and unlike rodents, hypoxia-induced PH in this species can lead to right heart failure. We, therefore, sought to examine the molecular and structural changes in the RV in calves with hypoxia-induced PH, hypothesizing that we could identify mechanisms underlying compensated physiological function in the face of developing severe PH. Calves were exposed to 14 days of environmental hypoxia (equivalent to 4,570 m/15,000 ft elevation, n = 29) or ambient normoxia (1,525 m/5,000 ft, n = 25). Cardiopulmonary function was evaluated by right heart catheterization and pressure volume loops. Molecular and cellular determinants of RV remodeling were analyzed by cDNA microarrays, RealTime PCR, proteomics, and immunochemistry. Hypoxic exposure induced robust PH, with increased RV contractile performance and preserved cardiac output, yet evidence of dysregulated RV-pulmonary artery mechanical coupling as seen in advanced disease. Analysis of gene expression revealed cellular processes associated with structural remodeling, cell signaling, and survival. We further identified specific clusters of gene expression associated with 1) hypertrophic gene expression and prosurvival mechanotransduction through YAP-TAZ signaling, 2) extracellular matrix (ECM) remodeling, 3) inflammatory cell activation, and 4) angiogenesis. A potential transcriptomic signature of cardiac fibroblasts in RV remodeling was detected, enriched in functions related to cell movement, tissue differentiation, and angiogenesis. Proteomic and immunohistochemical analysis confirmed RV myocyte hypertrophy, together with localization of ECM remodeling, inflammatory cell activation, and endothelial cell proliferation within the RV interstitium. In conclusion, hypoxia and hemodynamic load initiate coordinated processes of protective and compensatory RV remodeling to withstand the progression of PH.NEW & NOTEWORTHY Using a large animal model and employing a comprehensive approach integrating hemodynamic, transcriptomic, proteomic, and immunohistochemical analyses, we examined the early (2 wk) effects of severe PH on the RV. We observed that RV remodeling during PH progression represents a continuum of transcriptionally driven processes whereby cardiac myocytes, fibroblasts, endothelial cells, and proremodeling macrophages act to coordinately maintain physiological homeostasis and protect myocyte survival during chronic, severe, and progressive pressure overload.

The Effect of Hypoxia-inducible Factor Inhibition on the Phenotype of Fibroblasts in Human and Bovine Pulmonary Hypertension.

Hypoxia-inducible factor (HIF) has received much attention as a potential pulmonary hypertension (PH) treatment target because inhibition of HIF reduces the severity of established PH in rodent models. However, the limitations of small-animal models of PH in predicting the therapeutic effects of pharmacologic interventions in humans PH are well known. Therefore, we sought to interrogate the role of HIFs in driving the activated phenotype of PH cells from human and bovine vessels. We first established that pulmonary arteries (PAs) from human and bovine PH lungs exhibit markedly increased expression of direct HIF target genes (CA9, GLUT1, and NDRG1), as well as cytokines/chemokines (CCL2, CSF2, CXCL12, and IL6), growth factors (FGF1, FGF2, PDGFb, and TGFA), and apoptosis-resistance genes (BCL2, BCL2L1, and BIRC5). The expression of the genes found in the intact PAs was determined in endothelial cells, smooth muscle cells, and fibroblasts cultured from the PAs. The data showed that human and bovine pulmonary vascular fibroblasts from patients or animals with PH (termed PH-Fibs) were the cell type that exhibited the highest level and the most significant increases in the expression of cytokines/chemokines and growth factors. In addition, we found that human, but not bovine, PH-Fibs exhibit consistent misregulation of HIFα protein stability, reduced HIF1α protein hydroxylation, and increased expression of HIF target genes even in cells grown under normoxic conditions. However, whereas HIF inhibition reduced the expression of direct HIF target genes, it had no impact on other "persistently activated" genes. Thus, our study indicated that HIF inhibition alone is not sufficient to reverse the persistently activated phenotype of human and bovine PH-Fibs.

Novel EGFR-mutant mouse models of lung adenocarcinoma reveal adaptive immunity requirement for durable osimertinib response.

Lung cancers bearing oncogenically-mutated EGFR represent a significant fraction of lung adenocarcinomas (LUADs) for which EGFR-targeting tyrosine kinase inhibitors (TKIs) provide a highly effective therapeutic approach. However, these lung cancers eventually acquire resistance and undergo progression within a characteristically broad treatment duration range. Our previous study of EGFR mutant lung cancer patient biopsies highlighted the positive association of a TKI-induced interferon γ transcriptional response with increased time to treatment progression. To test the hypothesis that host immunity contributes to the TKI response, we developed novel genetically-engineered mouse models of EGFR mutant lung cancer bearing exon 19 deletions (del19) or the L860R missense mutation. Both oncogenic EGFR mouse models developed multifocal LUADs from which transplantable cancer cell lines sensitive to the EGFR-specific TKIs, gefitinib and osimertinib, were derived. When propagated orthotopically in the left lungs of syngeneic C57BL/6 mice, deep and durable shrinkage of the cell line-derived tumors was observed in response to daily treatment with osimertinib. By contrast, orthotopic tumors propagated in immune deficient nu/nu or Rag1-/- mice exhibited modest tumor shrinkage followed by rapid progression on continuous osimertinib treatment. Importantly, osimertinib treatment significantly increased intratumoral T cell content and decreased neutrophil content relative to diluent treatment. The findings provide strong evidence supporting the requirement for adaptive immunity in the durable therapeutic control of EGFR mutant lung cancer.

Secretion of IL-6 and IL-8 in the senescence of bone marrow mesenchymal stem cells is regulated by autophagy via FoxO3a.

Bone marrow mesenchymal stem cells (BMSCs) are widely used for therapeutic applications in tissue engineering and regenerative medicine. Nevertheless, the function of BMSCs is adversely affected by senescence. Thus, understanding the molecular mechanisms that contribute to BMSC senescence is critical for the development of BMSC-based tissue engineering and regenerative medicine. In this study, senescent BMSCs were characterized with >80 % of BMSCs stained positive for SA-ß-gal, increased expressions of senescence-related genes (p16INK4a and p21Waf1). These senescent characters were accompanied by elevated autophagic activity, up-regulation of interleukin 6 (IL-6), IL-8, and FoxO3a. Autophagic activity inhibition alleviated the senescent state with reduced levels of IL-6 and IL-8 during BMSC senescence. The enhanced autophagic activity upregulated the levels of IL-6 and IL-8 which is associated with up-regulation of FoxO3a, and knockdown of FoxO3a reduced IL-6 and IL-8 expression in senescent BMSCs. Therefore, this study indicated the pivotal role of autophagic activity in the expressions of IL-6 and IL-8 during BMSC senescence, which is regulated by FoxO3a.