Has2 Regulates the Development of Ovalbumin-Induced Airway Remodeling and Steroid Insensitivity in Mice

HAS2 is a member of the gene family encoding the hyaluronan synthase 2, which can generate high-molecular-weight hyaluronan (HMW-HA). Our previous study identified HAS2 as a candidate gene for increased susceptibility to adult asthma. However, whether HAS2 dysfunction affects airway remodeling and steroid insensitivity is still limited. Therefore, this study aimed to clarify the Has2 dysfunction, triggering severe airway remodeling and steroid insensitivity in a murine model of asthma. Has2 heterozygous-deficient (Has2+/−) mice and their wild-type littermates have been evaluated in a model of chronic ovalbumin (OVA) sensitization and challenge. Mice present a higher sensitivity to OVA and higher IL-17 release as well as eosinophilic infiltration. RNA sequencing demonstrated the downregulation of EIF2 signaling pathways, TGF-β signaling pathways, and heat shock proteins with Th17 bias in Has2+/−-OVA mice. The combined treatment with anti-IL-17A antibody and dexamethasone reduces steroid insensitivity in Has2+/−-OVA mice. Has2 attenuation worsens eosinophilic airway inflammation, airway remodeling, and steroid insensitivity. These data highlight that HAS2 and HMW-HA are important for controlling intractable eosinophilic airway inflammation and remodeling and could potentially be exploited for their therapeutic benefits in patients with asthma.

Hydrostatic pressure mimicking diurnal spinal movements maintains anabolic turnover in bovine nucleus pulposus cells in vitro

Treatment strategies for progressive intervertebral-disc degeneration often alleviate pain and other symptoms. With the goal of developing strategies to promote the regeneration of the nucleus pulposus (NP), the present study tried to identify the biological effects of hydrostatic (HP) and osmotic pressures on NP cells. The study hypothesis was that a repetitive regimen of cyclic HP followed by constant HP in high-osmolality medium would increase anabolic molecules in NP cells. Bovine NP cells/clusters were enclosed within semi-permeable membrane pouches and incubated under a regimen of cyclic HP for 2 d followed by constant HP for 1 d, repeated 6 times over 18 d. NP cells showed a significantly increased expression of anabolic genes over time: aggrecan, chondroitin sulfate N-acetylgalactosaminyltransferase 1, hyaluronan synthase 2, collagen type 2 (p < 0.05). In addition, the expression of catabolic or degenerative genes (matrix metalloproteinase 13, collagen type 1) and cellular characteristic genes (proliferating cell nucleic antigen, E-cadherin) was suppressed. The amount of sulfated glycosaminoglycan increased significantly at day 18 compared to day 3 (p < 0.01). Immunostaining revealed deposition of extracellular-matrix molecules and localization of other specific molecules corresponding to their genetic expression. An improved understanding of how cells respond to physicochemical stresses will help to better treat the degenerating disc using either cell- or gene-based therapies as well as other potential matrix-enhancing therapies. Efforts to apply these tissue-engineering and regenerative-medicine strategies will need to consider these important physicochemical stresses that may have a major impact on the survivability of such treatments.

Adipose tissue hyaluronan production improves systemic glucose homeostasis and primes adipocytes for CL 316,243-stimulated lipolysis

Plasma hyaluronan (HA) increases systemically in type 2 diabetes (T2D) and the HA synthesis inhibitor, 4-Methylumbelliferone, has been proposed to treat the disease. However, HA is also implicated in normal physiology. Therefore, we generated a Hyaluronan Synthase 2 transgenic mouse line, driven by a tet-response element promoter to understand the role of HA in systemic metabolism. To our surprise, adipocyte-specific overproduction of HA leads to smaller adipocytes and protects mice from high-fat-high-sucrose-diet-induced obesity and glucose intolerance. Adipocytes also have more free glycerol that can be released upon beta3 adrenergic stimulation. Improvements in glucose tolerance were not linked to increased plasma HA. Instead, an HA-driven systemic substrate redistribution and adipose tissue-liver crosstalk contributes to the systemic glucose improvements. In summary, we demonstrate an unexpected improvement in glucose metabolism as a consequence of HA overproduction in adipose tissue, which argues against the use of systemic HA synthesis inhibitors to treat obesity and T2D.

Differences in human placental mesenchymal stromal cells may impair vascular function in FGR

Placentae from pregnancies with fetal growth restriction (FGR) exhibit poor oxygen and nutrient exchange, in part due to impaired placental vascular development. Placental mesenchymal stromal cells (pMSCs) reside in a perivascular niche, where they may influence blood vessel formation/ function. However, the role of pMSCs in vascular dysfunction in FGR is unclear. To elucidate the mechanisms by which pMSCs may impact placental vascularisation we compared the transcriptomes of human pMSCs isolated from FGR (<5th centile) (n=7) and gestation-matched control placentae (n=9) using Affymetrix microarrays. At the transcriptome level there were no statistically significant differences between normal and FGR pMSCs, however several genes linked to vascular function exhibited notable fold changes, and thus the dataset was used as a hypothesis-generating tool for possible dysfunction in FGR. Genes/proteins of interest were followed up by real-time PCR and by immunohistochemistry. Gene expression of ADAMTS1 and FBLN2 (fibulin-2) were significantly upregulated, whilst HAS2 (hyaluronan synthase-2) was significantly downregulated, in pMSCs from FGR placentae (n=8) relative to controls (n=7, p<0.05 for all). At the protein level, significant differences in the level of fibulin-2 and hyaluronan synthase-2, but not ADAMTS1 were confirmed between pMSCs from FGR and control pregnancies by western blot. All three proteins demonstrated perivascular expression in third-trimester placentae. Fibulin-2 maintains vessel elasticity, and its increased expression in FGR pMSCs could help explain the increased distensibility of FGR blood vessels. ADAMTS1 and hyaluronan synthase-2 regulate angiogenesis, and their differential expression by FGR pMSCs may contribute to the impaired angiogenesis in these placentae.

Dysregulation of Pulmonary Responses in Severe COVID-19

Patients with coronavirus disease 2019 (COVID-19) predominantly have a respiratory tract infection with various symptoms and high mortality is associated with respiratory failure second to severe disease. The risk factors leading to severe disease remain unclear. Here, we reanalyzed a published single-cell RNA-Seq (scRNA-Seq) dataset and found that bronchoalveolar lavage fluid (BALF) of patients with severe disease compared to those with mild disease contained decreased TH17-type cells, decreased IFNA1-expressing cells with lower expression of toll-like receptor 7 (TLR7) and TLR8, increased IgA-expressing B cells, and increased hyperactive epithelial cells (and/or macrophages) expressing matrix metalloproteinases (MMPs), hyaluronan synthase 2 (HAS2), and plasminogen activator inhibitor-1 (PAI-1), which may together contribute to the pulmonary pathology in severe COVID-19. We propose IFN-I (and TLR7/TLR8) and PAI-1 as potential biomarkers to predict the susceptibility to severe COVID-19.

Lumican Inhibits In Vivo Melanoma Metastasis by Altering Matrix-Effectors and Invadopodia Markers

It was reported that lumican inhibits the activity of metalloproteinase MMP-14 and melanoma cell migration in vitro and in vivo. Moreover, Snail triggers epithelial-to-mesenchymal transition and the metastatic potential of cancer cells. Therefore, the aim of this study was to examine the effect of lumican on Mock and Snail overexpressing melanoma B16F1 cells in vivo. Lung metastasis was analyzed after intravenous injections of Mock-B16F1 and Snail-B16F1 cells in Lum+/+ and Lum−/− mice. At day 14, mice were sacrificed, and lungs were collected. The number of lung metastatic nodules was significantly higher in mice injected with Snail-B16F1 cells as compared to mice injected with Mock-B16F1 cells confirming the pro-metastatic effect of Snail. This effect was stronger in Lum−/− mice as compared to Lum+/+, suggesting that endogenous lumican of wild-type mice significantly inhibits metastasis to lungs. Scanning electron and confocal microscopy investigations demonstrated that lumican inhibits the development of elongated cancer cell phenotypes which are known to develop invadopodia releasing MMPs. Moreover, lumican was shown to affect the expression of cyclin D1, cortactin, vinculin, hyaluronan synthase 2, heparanase, MMP-14 and the phosphorylation of FAK, AKT, p130 Cas and GSK3α/β. Altogether, these data demonstrated that lumican significantly inhibits lung metastasis in vivo, as well as cell invasion in vitro, suggesting that a lumican-based strategy targeting Snail-induced metastasis could be useful for melanoma treatment.

PROGNOSTIC VALUES OF MIDKINE, SYNDECAN-1, HYALURONAN SYNTHASE-2, SESTRIN-1, LAMININ SUBUNIT ALPHA-4 AND FIBULIN-3 FOR MALIGNANT PLEURAL MESOTHELIOMA

IntroductionThe prognosis of malignant pleural mesothelioma (MPM) is poor with a limited survival time. In this study, we aimed to examine expression levels of genes selected from relevant literature and to utilize in silico methods to determine genes whose expression could reflect the prognosis of the patients with MPM by ex-vivo validation experimentsMaterial and methodsThe study group consisted of 54 MPM patients treated by chemotherapy. Expression of 6 genes; midkine (MDK), syndecan 1 (SDC1), hyaluronan synthase 2 (HAS2), sestrin 1 (SESN1), laminin subunit alpha 4 (LAMA4), and fibulin 3 (FBLN3) were examined by qPCR in tumor tissues. SESN1 and LAMA4 were identified using an in house R-based script “Unsupervised Survival Analysis Tool." MDK, SDC1, HAS2, and FBLN3 were selected from current literature. We used two housekeeping genes; glucose-6-phosphate dehydrogenase and TATA-box binding protein as controls.ResultsOf the patients, 43 (79.6%) had epithelioid mesothelioma. The median survival for all patients was 10 (±1.2 SE) months (CI 95%; 7.7-12.3). In multivariate analyses, MDK (p=0.007), HAS2 (p=0.008) and SESN1 (p=0.014) expression were related with survival time in whole group. In epithelioid type MPM patients, MDK (p=0.014), FBLN3 (p=0.029), HAS2 (p=0.014) and SESN1 (p=0.045) expression was related with survival time by multivariate analyses.ConclusionsHigh HAS2 and SESN1 expressions and low MDK are potential biomarkers of good prognosis in MPM. High HAS2 and SESN1 expression and low MDK and FBLN3 can also be utilized as biomarkers of good prognosis for epithelioid MPM. Those results should be further investigated in sera, plasma, and pleural effusions

Stromal NRG1 in luminal breast cancer defines pro-fibrotic and migratory cancer-associated fibroblasts

HER3 is highly expressed in luminal breast cancer subtypes. Its activation by NRG1 promotes activation of AKT and ERK1/2, contributing to tumour progression and therapy resistance. HER3-targeting agents that block this activation, are currently under phase 1/2 clinical studies, and although they have shown favorable tolerability, their activity as a single agent has proven to be limited. Here we show that phosphorylation and activation of HER3 in luminal breast cancer cells occurs in a paracrine manner and is mediated by NRG1 expressed by cancer-associated fibroblasts (CAFs). Moreover, we uncover a HER3-independent NRG1 signaling in CAFs that results in the induction of a strong migratory and pro-fibrotic phenotype, describing a subtype of CAFs with elevated expression of NRG1 and an associated transcriptomic profile that determines their functional properties. Finally, we identified Hyaluronan Synthase 2 (HAS2), a targetable molecule strongly correlated with NRG1, as an attractive player supporting NRG1 signaling in CAFs.

The Secreted Protein C10orf118 Is a New Regulator of Hyaluronan Synthesis Involved in Tumour-Stroma Cross-Talk

Interaction between cancer cells and their microenvironment is central in defining the fate of cancer development. Tumour cells secrete signals (cytokines, chemokines, growth factors) that modify the surrounding area, while the niche supplies structures and activities necessary for tumour maintenance and growth. Hyaluronan (HA) is a glycosaminoglycan that constitute cancer cell niche and is known to influence tumour functions such as proliferation, migration and neoangiogenesis. The knowledge of the factors regulating HA synthesis and size is crucial in understanding the mechanisms sustaining tumour development. Here we show that a yet uncharacterized protein secreted by breast tumour cell lines, named c10orf118 (accession number NM_018017 in NCBI/BLAST, and Q7z3E2 according to the Uniprot identifier), with a predicted length of 898 amino acids, can induce the secretion of HA by stromal fibroblasts through the up-regulation of the hyaluronan synthase 2 gene (HAS2). Intracellularly, this protein is localized in the Golgi apparatus with a possible role in vesicle maturation and transport. The expression of c10orf118 was verified in breast cancer patient specimens and was found to be associated with the presence of estrogen receptor that characterizes a good patient survival. We suggest c10orf118 as a new player that influences the HA amount in breast cancer microenvironment and is associated with low aggressiveness of cancer.