Efficacy and pharmacokinetics of subcutaneous exendin (9-39) in patients with post-bariatric hypoglycaemia.

AIM: To evaluate the efficacy, pharmacokinetic (PK) profile and tolerability of subcutaneous (s.c.). exendin 9-39 (Ex-9) injection in patients with post-bariatric hypoglycaemia (PBH). METHODS: Nine women who had recurrent symptomatic hypoglycaemia after undergoing Roux-en-Y gastric bypass were enrolled in this 2-part, single-blind, single-ascending-dose study. In Part 1, a single participant underwent equimolar low-dose intravenous (i.v.) vs s.c. Ex-9 administration; in Part 2, 8 participants were administered single ascending doses of s.c. Ex-9 during an oral glucose tolerance test (OGTT). Glycaemic, hormonal, PK and symptomatic responses were compared with those obtained during the baseline OGTT. RESULTS: Although an exposure-response relationship was observed, all doses effectively prevented hyperinsulinaemic hypoglycaemia and improved associated symptoms. On average, the postprandial glucose nadir was increased by 66%, peak insulin was reduced by 57%, and neuroglycopenic symptoms were reduced by 80%. All doses were well tolerated with no treatment-emergent adverse events observed. CONCLUSIONS: Injection s.c. of Ex-9 appears to represent a safe, effective and targeted therapeutic approach for treatment of PBH. Further investigation involving multiple doses with chronic dosing is warranted.

Critical role for GLP-1 in symptomatic post-bariatric hypoglycaemia.

AIMS/HYPOTHESIS: Post-bariatric hypoglycaemia (PBH) is a rare, but severe, metabolic disorder arising months to years after bariatric surgery. It is characterised by symptomatic postprandial hypoglycaemia, with inappropriately elevated insulin concentrations. The relative contribution of exaggerated incretin hormone signalling to dysregulated insulin secretion and symptomatic hypoglycaemia is a subject of ongoing inquiry. This study was designed to test the hypothesis that PBH and associated symptoms are primarily mediated by glucagon-like peptide-1 (GLP-1). METHODS: We conducted a double-blinded crossover study wherein eight participants with confirmed PBH were assigned in random order to intravenous infusion of the GLP-1 receptor (GLP-1r) antagonist. Exendin (9-39) (Ex-9), or placebo during an OGTT on two separate days at the Stanford University Clinical and Translational Research Unit. Metabolic, symptomatic and pharmacokinetic variables were evaluated. Results were compared with a cohort of BMI- and glucose-matched non-surgical controls (NSCs). RESULTS: Infusion of Ex-9 decreased the time to peak glucose and rate of glucose decline during OGTT, and raised the postprandial nadir by over 70%, normalising it relative to NSCs and preventing hypoglycaemia in all PBH participants. Insulin AUC and secretion rate decreased by 57% and 71% respectively, and peak postprandial insulin was normalised relative to NSCs. Autonomic and neuroglycopenic symptoms were significantly reduced during Ex-9 infusion. CONCLUSIONS/INTERPRETATION: GLP-1r blockade prevented hypoglycaemia in 100% of individuals, normalised beta cell function and reversed neuroglycopenic symptoms, supporting the conclusion that GLP-1 plays a primary role in mediating hyperinsulinaemic hypoglycaemia in PBH. Competitive antagonism at the GLP-1r merits consideration as a therapeutic strategy. TRIAL REGISTRATION: ClinicalTrials.gov NCT02550145.

The GLP-1 receptor agonists exendin-4 and liraglutide alleviate oxidative stress and cognitive and micturition deficits induced by middle cerebral artery occlusion in diabetic mice.

BACKGROUND: Glucagon-like peptide 1 (GLP-1) analogs protect a variety of cell types against oxidative damage and vascular and neuronal injury via binding to GLP-1 receptors. This study aimed to investigate the effects of the GLP-1 analogs exendin-4 and liraglutide on cerebral blood flow, reactive oxygen species production, expression of oxidative stress-related proteins, cognition, and pelvic sympathetic nerve-mediated bladder contraction after middle cerebral artery occlusion (MCAO) injury in the db/db mouse model of diabetes. RESULTS: Sixty minutes of MCAO increased blood and brain reactive oxygen species counts in male db/db mice, as revealed by dihydroethidium staining. MCAO also increased nuclear factor-κB and intercellular adhesion molecule-1 expression and decreased cerebral microcirculation. These effects were attenuated by treatment with exendin-4 or liraglutide. MCAO did not affect basal levels of phosphorylated Akt (p-Akt) or endothelial nitric oxide synthase (p-eNOS); however, exendin-4 and liraglutide treatments significantly enhanced p-Akt and p-eNOS levels, indicating activation of the p-Akt/p-eNOS signaling pathway. MCAO-induced motor and cognitive deficits and micturition dysfunction, indicated by reduced pelvic nerve-mediated voiding contractions and increased nonvoiding contractions, were also partially attenuated by exendin-4 treatment. CONCLUSIONS: The above data indicate that treatment with GLP-1 agonists exerts protective effects against oxidative, inflammatory, and apoptotic damage in brain areas that control parasympathetic/pelvic nerve-mediated voiding contractions and cognitive and motor behaviors in a diabetic mouse model.

In vivo 2H2O administration reveals impaired triglyceride storage in adipose tissue of insulin-resistant humans.

Indirect evidence suggests that impaired triglyceride storage in the subcutaneous fat depot contributes to the development of insulin resistance via lipotoxicity. We directly tested this hypothesis by measuring, in vivo, TG synthesis, de novo lipogenesis (DNL), adipocyte proliferation, and insulin suppression of lipolysis in subcutaneous adipose tissue of BMI-matched individuals classified as insulin resistant (IR) or insulin sensitive (IS). Nondiabetic, moderately obese subjects with BMI 25-35 kg/m(2), classified as IR or IS by the modified insulin suppression test, consumed deuterated water ((2)H2O) for 4 weeks. Deuterium incorporation into glycerol, palmitate, and DNA indicated TG synthesis, DNL, and adipocyte proliferation, respectively. Net TG synthesis and DNL in adipose cells were significantly lower in IR as compared with IS subjects, whereas adipocyte proliferation did not differ significantly. Plasma FFAs measured during an insulin suppression test were 2.5-fold higher in IR subjects, indicating resistance to insulin suppression of lipolysis. Adipose TG synthesis correlated directly with DNL but not with proliferation. These results provide direct in vivo evidence for impaired TG storage in subcutaneous adipose tissue of IR as compared with IS. Relative inability to store TG in the subcutaneous depot may represent a mechanism contributing to the development of insulin resistance in the setting of obesity.

The receptor CD44 is associated with systemic insulin resistance and proinflammatory macrophages in human adipose tissue.

AIMS/HYPOTHESIS: Proinflammatory immune cell infiltration in human adipose tissue is associated with the development of insulin resistance. We previously identified, via a gene expression-based genome-wide association study, the cell-surface immune cell receptor CD44 as a functionally important gene associated with type 2 diabetes. We then showed that, compared with controls, Cd44 knockout mice were protected from insulin resistance and adipose tissue inflammation during diet-induced obesity. We thus sought to test whether CD44 is associated with adipose tissue inflammation and insulin resistance in humans. METHODS: Participants included 58 healthy, overweight/moderately obese white adults who met predetermined criteria for insulin resistance or insulin sensitivity based on the modified insulin-suppression test. Serum was collected from 43 participants to measure circulating concentrations of CD44. Subcutaneous adipose tissue was obtained from 17 participants to compare CD44, its ligand osteopontin (OPN, also known as SPP1) and pro-inflammatory gene expression. CD44 expression on adipose tissue macrophage (ATM) surfaces was determined by flow cytometry. RESULTS: Serum CD44 concentrations were significantly increased in insulin-resistant (IR) participants. CD44 gene expression in subcutaneous adipose tissue was threefold higher in the IR subgroup. The expression of OPN, CD68 and IL6 was also significantly elevated in IR individuals. CD44 gene expression correlated significantly with CD68 and IL6 expression. CD44 density on ATMs was associated with proinflammatory M1 polarisation. CONCLUSIONS/INTERPRETATION: CD44 and OPN in human adipose tissue are associated with localised inflammation and systemic insulin resistance. This receptor-ligand pair is worthy of further research as a potentially modifiable contributor to human insulin resistance and type 2 diabetes.

T-cell profile in adipose tissue is associated with insulin resistance and systemic inflammation in humans.

OBJECTIVE: The biological mechanisms linking obesity to insulin resistance have not been fully elucidated. We have shown that insulin resistance or glucose intolerance in diet-induced obese mice is related to a shift in the ratio of pro- and anti-inflammatory T cells in adipose tissue. We sought to test the hypothesis that the balance of T-cell phenotypes would be similarly related to insulin resistance in human obesity. APPROACH AND RESULTS: Healthy overweight or obese human subjects underwent adipose-tissue biopsies and quantification of insulin-mediated glucose disposal by the modified insulin suppression test. T-cell subsets were quantified by flow cytometry in visceral (VAT) and subcutaneous adipose tissue (SAT). Results showed that CD4 and CD8 T cells infiltrate both depots, with proinflammatory T-helper (Th)-1, Th17, and CD8 T cells, significantly more frequent in VAT as compared with SAT. T-cell profiles in SAT and VAT correlated significantly with one another and with peripheral blood. Th1 frequency in SAT and VAT correlated directly, whereas Th2 frequency in VAT correlated inversely, with plasma high-sensitivity C-reactive protein concentrations. Th2 in both depots and peripheral blood was inversely associated with systemic insulin resistance. Furthermore, Th1 in SAT correlated with plasma interleukin-6. Relative expression of associated cytokines, measured by real-time polymerase chain reaction, reflected flow cytometry results. Most notably, adipose tissue expression of anti-inflammatory interleukin-10 was inversely associated with insulin resistance. CONCLUSIONS: CD4 and CD8 T cells populate human adipose tissue and the relative frequency of Th1 and Th2 are highly associated with systemic inflammation and insulin resistance. These findings point to the adaptive immune system as a potential mediator between obesity and insulin resistance or inflammation. Identification of antigenic stimuli in adipose tissue may yield novel targets for treatment of obesity-associated metabolic disease.

Effects of Aloe Gel on Lactating Women with Nipple Trauma.

Background: To investigate the efficacy of aloe gel in reducing pain and promoting wound healing in postpartum women with nipple trauma. Method: There were 80 postpartum women who took part in this study having developed nipple trauma during breastfeeding in the obstetrics department of a tertiary grade A hospital in Suzhou from January to December 2021. Postpartum women with nipple trauma whose hospital bed numbers ranged between 15 and 33 were included in the test group, whereas those whose hospital bed numbers ranged between 35 and 53 were included in the control group. Both groups received health education and breastfeeding guidance. The control group applied lanolin cream to their nipple trauma, whereas the test group used aloe gel. We used a nipple trauma severity assessment table to determine the severity of nipple trauma in lactating women and a Visual Analogue Scale (VAS) to determine the level of nipple pain and referred to the Traditional Chinese Medicine Standard for Diagnosis and Therapeutic Efficacy for Diseases and Syndromes to determine the healing time of their wounds. Results: The test group scored 3.70 ± 1.24 and 1.65 ± 0.74 points on the VAS on the first and third days following the intervention, whereas the control group scored 4.30 ± 0.94 and 2.23 ± 1.07 points, respectively. It took 3.75 ± 1.08 days and 4.45 ± 1.15 days for the nipple pain to completely disappear in the test group and the control group, respectively. The healing period for nipple trauma was 5.28 ± 1.26 days for the test group and 6.03 ± 1.61 days for the control group. All of the aforementioned distinctions were statistically significant (p < 0.05). Conclusions: Aloe gel can significantly alleviate the pain associated with nipple trauma in lactating women, accelerate wound healing, and reduce the duration of nipple trauma.

Inhibiting 5-hydroxytryptamine receptor 3 alleviates pathological changes of a mouse model of Alzheimer's disease.

Extracellular amyloid beta (Aß) plaques are main pathological feature of Alzheimer's disease. However, the specific type of neurons that produce Aß peptides in the initial stage of Alzheimer's disease are unknown. In this study, we found that 5-hydroxytryptamin receptor 3A subunit (HTR3A) was highly expressed in the brain tissue of transgenic amyloid precursor protein and presenilin-1 mice (an Alzheimer's disease model) and patients with Alzheimer's disease. To investigate whether HTR3A-positive interneurons are associated with the production of Aß plaques, we performed double immunostaining and found that HTR3A-positive interneurons were clustered around Aß plaques in the mouse model. Some amyloid precursor protein-positive or ß-site amyloid precursor protein cleaving enzyme-1-positive neurites near Aß plaques were co-localized with HTR3A interneurons. These results suggest that HTR3A -positive interneurons may partially contribute to the generation of Aß peptides. We treated 5.0-5.5-month-old model mice with tropisetron, a HTR3 antagonist, for 8 consecutive weeks. We found that the cognitive deficit of mice was partially reversed, Aß plaques and neuroinflammation were remarkably reduced, the expression of HTR3 was remarkably decreased and the calcineurin/nuclear factor of activated T-cell 4 signaling pathway was inhibited in treated model mice. These findings suggest that HTR3A interneurons partly contribute to generation of Aß peptide at the initial stage of Alzheimer's disease and inhibiting HTR3 partly reverses the pathological changes of Alzheimer's disease.

Neuroplastin 65 deficiency reduces amyloid plaque formation and cognitive deficits in an Alzheimer's disease mouse model.

Introduction: Alzheimer's disease (AD) is characterized by increasing cognitive dysfunction, progressive cerebral amyloid beta (Aß) deposition, and neurofibrillary tangle aggregation. However, the molecular mechanisms of AD pathologies have not been completely understood. As synaptic glycoprotein neuroplastin 65 (NP65) is related with synaptic plasticity and complex molecular events underlying learning and memory, we hypothesized that NP65 would be involved in cognitive dysfunction and Aß plaque formation of AD. For this purpose, we examined the role of NP65 in the transgenic amyloid precursor protein (APP)/presenilin 1 (PS1) mouse model of AD. Methods: Neuroplastin 65-knockout (NP65-/-) mice crossed with APP/PS1 mice to get the NP65-deficient APP/PS1 mice. In the present study, a separate cohort of NP65-deficient APP/PS1 mice were used. First, the cognitive behaviors of NP65-deficient APP/PS1 mice were assessed. Then, Aß plaque burden and Aß levels in NP65-deficient APP/PS1 mice were measured by immunostaining and western blot as well as ELISA. Thirdly, immunostaining and western blot were used to evaluate the glial response and neuroinflammation. Finally, protein levels of 5-hydroxytryptamin (serotonin) receptor 3A and synaptic proteins and neurons were measured. Results: We found that loss of NP65 alleviated the cognitive deficits of APP/PS1 mice. In addition, Aß plaque burden and Aß levels were significantly reduced in NP65-deficient APP/PS1 mice compared with control animals. NP65-loss in APP/PS1 mice resulted in a decrease in glial activation and the levels of pro- and anti-inflammatory cytokines (IL-1ß, TNF-α, and IL-4) as well as protective matrix YM-1 and Arg-1, but had no effect on microglial phenotype. Moreover, NP65 deficiency significantly reversed the increase in 5-hydroxytryptamine (serotonin) receptor 3A (Htr3A) expression levels in the hippocampus of APP/PS1 mice. Discussion: These findings identify a previously unrecognized role of NP65 in cognitive deficits and Aß formation of APP/PS1 mice, and suggest that NP65 may serve as a potential therapeutic target for AD.

CD8+ T cell-intrinsic IL-6 signaling promotes resistance to anti-PD-L1 immunotherapy.

Although immune checkpoint inhibitors (ICIs) are established as effective cancer therapies, overcoming therapeutic resistance remains a critical challenge. Here we identify interleukin 6 (IL-6) as a correlate of poor response to atezolizumab (anti-PD-L1) in large clinical trials of advanced kidney, breast, and bladder cancers. In pre-clinical models, combined blockade of PD-L1 and the IL-6 receptor (IL6R) causes synergistic regression of large established tumors and substantially improves anti-tumor CD8+ cytotoxic T lymphocyte (CTL) responses compared with anti-PD-L1 alone. Circulating CTLs from cancer patients with high plasma IL-6 display a repressed functional profile based on single-cell RNA sequencing, and IL-6-STAT3 signaling inhibits classical cytotoxic differentiation of CTLs in vitro. In tumor-bearing mice, CTL-specific IL6R deficiency is sufficient to improve anti-PD-L1 activity. Thus, based on both clinical and experimental evidence, agents targeting IL-6 signaling are plausible partners for combination with ICIs in cancer patients.

Hormone-sensitive lipase modulates adipose metabolism through PPARγ.

Hormone-sensitive lipase (HSL) is rate limiting for diacylglycerol and cholesteryl ester hydrolysis in adipose tissue and essential for complete hormone-stimulated lipolysis. Gene expression profiling in HSL-/- mice suggests that HSL is important for modulating adipogenesis and adipose metabolism. To test whether HSL is required for the supply of intrinsic ligands for PPARγ for normal adipose differentiation, HSL-/- and wild-type (WT) littermates were fed normal chow (NC) and high-fat (HF) diets supplemented with or without rosiglitazone (200 mg/kg) for 16 weeks. Results show that supplementing rosiglitazone to an NC diet completely normalized the decreased body weight and adipose depots in HSL-/- mice. Additionally, rosiglitazone resulted in similar serum glucose, total cholesterol, FFA, and adiponectin values in WT and HSL-/- mice. Furthermore, rosiglitazone normalized the expression of genes involved in adipocyte differentiation, markers of adipocyte differentiation, and enzymes involved in triacylglycerol synthesis and metabolism, and cholesteryl ester homeostasis, in HSL-/- mice. Supplementing rosiglitazone to an HF diet resulted in improved glucose tolerance in both WT and HSL-/- animals and also partial normalization in HSL-/- mice of abnormal WAT gene expression, serum chemistries, organ and body weight changes. In vitro studies showed that adipocytes from WT animals can provide ligands for activation of PPARγ and that activation is further boosted following lipolytic stimulation, whereas adipocytes from HSL-/- mice displayed attenuated activation of PPARγ, with no change following lipolytic stimulation. These results suggest that one of the mechanisms by which HSL modulates adipose metabolism is by providing intrinsic ligands or pro-ligands for PPARγ.

Hepatoma-derived growth factor stimulates podosome rosettes formation in NIH/3T3 cells through the activation of phosphatidylinositol 3-kinase/Akt pathway.

Hepatoma-derived growth factor (HDGF) stimulates the migration, invasion and metastasis in several types of cancer cells. However, the mechanism underlying HDGF-stimulated migration remains unclear. In this study, we investigated the influence of HDGF on cytoskeleton remodeling and phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway in non-transformed NIH/3T3 cells. Exogenous HDGF promoted the migration and the formation of dorsal ruffles and podosome rosettes. Besides, HDGF supply increased the PI3K expression and Akt phosphorylation in dose- and time-dependent manners. Application of LY294002, a PI3K inhibitor, attenuated the HDGF-induced migration, dorsal ruffles and podosome rosettes formation. Consistently, the HDGF-overexpressing NIH/3T3 transfectants exhibited significantly increased motility and elevated PI3K/Akt activities, which were repressed by LY294002 or adenovirus-mediated overexpression of endogenous PI3K antagonist, PTEN. In summary, HDGF elicits the activation of PI3K/Akt signaling cascade, thereby promoting cytoskeleton remodeling to stimulate cellular migration.

Hormone-sensitive lipase-knockout mice maintain high bone density during aging.

We tested the hypothesis that the actions of hormone-sensitive lipase (HSL) affect the microenvironment of the bone marrow and that removal of HSL function by gene deletion maintains high bone mass in aging mice. We compared littermate control wild-type (WT) and HSL(-/-) mice during aging for changes in serum biochemical values, trabecular bone density using micro-computed tomography, bone histomorphometry, and characteristics of primary bone marrow cells and preosteoblasts. There is a regulated expression of HSL and genes involved in lipid metabolism in the bone marrow during aging. HSL(-/-) mice have increased serum levels of insulin and osteocalcin with decreased leptin levels. Compared with the marked adipocyte infiltration in WT bone marrow (65% by area) at 14 mo, HSL(-/-) mice have fewer (16%, P<0.05) and smaller adipocytes in bone marrow. While peak bone density is similar, HSL(-/-) mice maintain a higher bone density (bone volume/total volume 6.1%) with age than WT mice (2.6%, P<0.05). Primary osteoblasts from HSL(-/-) mice show increased growth rates and higher osteogenic potential, manifested by increased expression of Runx2 (3.5-fold, P<0.05) and osteocalcin (4-fold, P<0.05). The absence of HSL directs cells within the bone marrow toward osteoblast differentiation and favors the maintenance of bone density with aging.

Characterization of age-related gene expression profiling in bone marrow and epididymal adipocytes.

BACKGROUND: While an increase in bone marrow adiposity is associated with age-related bone disease, the function of bone marrow adipocytes has not been studied. The aim of this study was to characterize and compare the age-related gene expression profiles in bone marrow adipocytes and epididymal adipocytes. RESULTS: A total of 3918 (13.7%) genes were differentially expressed in bone marrow adipocytes compared to epididymal adipocytes. Bone marrow adipocytes revealed a distinct gene profile with low expression of adipocyte-specific genes peroxisome proliferator-activated receptor gamma (PPARγ), fatty acid binding protein 4 (FABP4), perilipin (Plin1), adipsin (CFD) and high expression of genes associated with early adipocyte differentiation (CCAAT/enhancer binding protein beta (C/EBPß), regulator of G-protein signaling 2 (RGS2). In addition, a number of genes including secreted frizzled related protein 4 (SFRP4), tumor necrosis factor α (TNFα), transforming growth factor beta 1(TGFß1), G-protein coupled receptor 109A (GPR109A) and interleukin 6 (IL-6), that could affect adipose-derived signaling to bone are markedly increased in bone marrow adipocytes. Age had a substantial effect on genes associated with mitochondria function and inflammation in bone marrow adipocytes. Twenty seven genes were significantly changed with age in both adipocyte depots. Among these genes, IL6 and GPR109A were significantly reduced with age in both adipocyte depots. CONCLUSIONS: Overall, gene profiling reveals a unique phenotype for primary bone marrow adipocytes characterized by low adipose-specific gene expression and high expression of inflammatory response genes. Bone marrow and epididymal adipocytes share a common pathway in response to aging in mice, but age has a greater impact on global gene expression in epididymal than in bone marrow adipocytes. Genes that are differentially expressed at greater levels in the bone marrow are highly regulated with age.

Effects of primary microglia and astrocytes on neural stem cells in in vitro and in vivo models of ischemic stroke.

Transplantation of neural stem cells (NSCs) can protect neurons in animal stroke models; however, their low rates of survival and neuronal differentiation limit their clinical application. Glial niches, an important location of neural stem cells, regulate survival, proliferation and differentiation of neural stem cells. However, the effects of activated glial cells on neural stem cells remain unclear. In the present study, we explored the effects of activated astrocytes and microglia on neural stem cells in vitro stroke models. We also investigated the effects of combined transplantation of neural stem cells and glial cells after stroke in rats. In a Transwell co-culture system, primary cultured astrocytes, microglia or mixed glial cells were exposed to glutamate or H2O2 and then seeded in the upper inserts, while primary neural stem cells were seeded in the lower uncoated wells and cultured for 7 days. Our results showed that microglia were conducive to neurosphere formation and had no effects on apoptosis within neurospheres, while astrocytes and mixed glial cells were conducive to neurosphere differentiation and reduced apoptosis within neurospheres, regardless of their pretreatment. In contrast, microglia and astrocytes induced neuronal differentiation of neural stem cells in differentiation medium, regardless of their pretreatment, with an exception of astrocytes pretreated with H2O2. Rat models of ischemic stroke were established by occlusion of the middle cerebral artery. Three days later, 5 × 105 neural stem cells with microglia or astrocytes were injected into the right lateral ventricle. Neural stem cell/astrocyte-treated rats displayed better improvement of neurological deficits than neural stem cell only-treated rats at 4 days after cell transplantation. Moreover, neural stem cell/microglia-, and neural stem cell/astrocyte-treated rats showed a significant decrease in ischemic volume compared with neural stem cell-treated rats. These findings indicate that microglia and astrocytes exert different effects on neural stem cells, and that co-transplantation of neural stem cells and astrocytes is more conducive to the recovery of neurological impairment in rats with ischemic stroke. The study was approved by the Animal Ethics Committee of Tongji University School of Medicine, China (approval No. 2010-TJAA08220401) in 2010.

Molecular determinants of response to PD-L1 blockade across tumor types.

Immune checkpoint inhibitors targeting the PD-1/PD-L1 axis lead to durable clinical responses in subsets of cancer patients across multiple indications, including non-small cell lung cancer (NSCLC), urothelial carcinoma (UC) and renal cell carcinoma (RCC). Herein, we complement PD-L1 immunohistochemistry (IHC) and tumor mutation burden (TMB) with RNA-seq in 366 patients to identify unifying and indication-specific molecular profiles that can predict response to checkpoint blockade across these tumor types. Multiple machine learning approaches failed to identify a baseline transcriptional signature highly predictive of response across these indications. Signatures described previously for immune checkpoint inhibitors also failed to validate. At the pathway level, significant heterogeneity is observed between indications, in particular within the PD-L1+ tumors. mUC and NSCLC are molecularly aligned, with cell cycle and DNA damage repair genes associated with response in PD-L1- tumors. At the gene level, the CDK4/6 inhibitor CDKN2A is identified as a significant transcriptional correlate of response, highlighting the association of non-immune pathways to the outcome of checkpoint blockade. This cross-indication analysis reveals molecular heterogeneity between mUC, NSCLC and RCC tumors, suggesting that indication-specific molecular approaches should be prioritized to formulate treatment strategies.

Adipocytes decrease Runx2 expression in osteoblastic cells: roles of PPARγ and adiponectin.

The mechanisms through which bone marrow adipocytes might influence differentiation and function of osteoblasts are not completely understood. To investigate the direct effects of bone marrow fat cells on osteoblast function, an ex vivo co-culture system was utilized comprising either primary fat cells or differentiated 3T3-L1 adipocytes and osteoblastic cells on transwells. In co-culture, both adipocytes and osteoblastic cells were differentiated into adipocytes or osteoblasts, respectively, before culturing on transwells. Co-culture with either primary fat cells or fully differentiated 3T3-L1 adipocytes significantly decreased mRNA and protein expression of runt-related transcription factor 2 (Runx2) in osteoblastic cells. An increase in mRNA and protein expression of peroxisome proliferator-activated receptor γ (PPARγ) occurred concomitantly with the reduction of Runx2 expression. Adiponectin concentration was increased in the media by co-culture. In addition, co-culture with conditioned media from fat cells increased PPARγ promoter activity and decreased Runx2 promoter activity. Knockdown of PPARγ or adiponectin receptor 1 in osteoblastic cells by siRNA prevented the down-regulation of mRNA expression of Runx2 in osteoblastic cells cultured with fully differentiated 3T3-L1 cells. Furthermore, co-transfection with PPARγ decreased Runx2 promoter activity. A marker of osteogenesis, alkaline phosphatase activity in osteoblastic cells was significantly decreased by co-culture. Annexin V/propidium iodide staining showed that co-culture did not induce apoptosis in osteoblastic cells. Thus, we conclude that adipocytes modulate key metabolic functions of osteoblasts through the release of secretory products. PPARγ plays a key role in mediating the effects of adipocytes on osteoblasts.

Molecular Subsets in Renal Cancer Determine Outcome to Checkpoint and Angiogenesis Blockade.

Integrated multi-omics evaluation of 823 tumors from advanced renal cell carcinoma (RCC) patients identifies molecular subsets associated with differential clinical outcomes to angiogenesis blockade alone or with a checkpoint inhibitor. Unsupervised transcriptomic analysis reveals seven molecular subsets with distinct angiogenesis, immune, cell-cycle, metabolism, and stromal programs. While sunitinib and atezolizumab + bevacizumab are effective in subsets with high angiogenesis, atezolizumab + bevacizumab improves clinical benefit in tumors with high T-effector and/or cell-cycle transcription. Somatic mutations in PBRM1 and KDM5C associate with high angiogenesis and AMPK/fatty acid oxidation gene expression, while CDKN2A/B and TP53 alterations associate with increased cell-cycle and anabolic metabolism. Sarcomatoid tumors exhibit lower prevalence of PBRM1 mutations and angiogenesis markers, frequent CDKN2A/B alterations, and increased PD-L1 expression. These findings can be applied to molecularly stratify patients, explain improved outcomes of sarcomatoid tumors to checkpoint blockade versus antiangiogenics alone, and develop personalized therapies in RCC and other indications.

High systemic and tumor-associated IL-8 correlates with reduced clinical benefit of PD-L1 blockade.

Although elevated plasma interleukin-8 (pIL-8) has been associated with poor outcome to immune checkpoint blockade 1, this has not been comprehensively evaluated in large randomized studies. Here we analyzed circulating pIL-8 and IL8 gene expression in peripheral blood mononuclear cells and tumors of patients treated with atezolizumab (anti-PD-L1 monoclonal antibody) from multiple randomized trials representing 1,445 patients with metastatic urothelial carcinoma (mUC) and metastatic renal cell carcinoma. High levels of IL-8 in plasma, peripheral blood mononuclear cells and tumors were associated with decreased efficacy of atezolizumab in patients with mUC and metastatic renal cell carcinoma, even in tumors that were classically CD8+ T cell inflamed. Low baseline pIL-8 in patients with mUC was associated with increased response to atezolizumab and chemotherapy. Patients with mUC who experienced on-treatment decreases in pIL-8 exhibited improved overall survival when treated with atezolizumab but not with chemotherapy. Single-cell RNA sequencing of the immune compartment showed that IL8 is primarily expressed in circulating and intratumoral myeloid cells and that high IL8 expression is associated with downregulation of the antigen-presentation machinery. Therapies that can reverse the impacts of IL-8-mediated myeloid inflammation will be essential for improving outcomes of patients treated with immune checkpoint inhibitors.

[Personality and occupation stress of workers in processing line in Guangzhou].

OBJECTIVE: To study relationships between personality and occupational stress of workers in processing line in Guangzhou. METHODS: 452 workers in processing line in Guangzhou were investigated by the Eysenck Personality Questionnaire (EPQ) and Occupational Stress Indicator (OSI). RESULTS: N scores and P scores of EPQ were positively correlated with the occupational stress and the poor physical fitness status (r value was 0.196, 0.128, 0.202, 0.218, respectively) (P < 0.01). N scores of EPQ were positively correlated with the type A behavior (r = 0.164) and negatively correlated with the mental well-being level (r = -0.196, P < 0.01). P scores of EPQ were negative correlated with the sense of job satisfaction and the stress coping ability(r value was -0.146, -0.140, respectively) (P < 0.01). E scores and L scores of EPQ were negatively correlated with the type A behavior (r value was -0.159, -0.138, respectively) (P < 0.01). E scores of EPQ were positively correlated with the stress coping ability (r = 0.176, P < 0.01). L scores of EPQ were negatively correlated with the occupational stress, the poor physical fitness status and the explanation to vicinity event (r value was -0.139, -0.140, -0.111, respectively) (P < 0.01, P < 0.05). CONCLUSION: The personality of neuroticism, psychoticism and introversion may be related to the occupational stress, the sense of job satisfaction and unhealthy mental reaction.