Pro-angiogenic Factors Research Articles

A randomized, controlled, phase 2 trial of nivolumab plus standard-dose or low-dose bevacizumab for recurrent glioblastoma (NAVAL).

2072 Background: PD-1/PD-L1 inhibition has demonstrated limited efficacy for recurrent glioblastoma (rGBM) across prior RCTs. VEGF, a proangiogenic factor upregulated in rGBM, contributes to tumor-associated immunosuppression. Preclinical and observational clinical data indicate a potential dose-dependent effect of anti-VEGF therapy on immune response. This RCT evaluated anti-PD-1 plus anti-VEGF therapy for rGBM. Methods: NAVAL was a multicenter, open label, phase 2 RCT that enrolled 90 patients (≥18 years) with GBM at first recurrence. Participants received nivolumab (240 mg IV Q2 weeks), and were 1:1 randomized to concurrent 10 mg/kg bevacizumab ( standard-dose arm) or 3 mg/kg bevacizumab ( low-dose arm) administered IV biweekly. Stratification factors included age, performance status, extent of resection, and MGMT methylation status. Primary endpoint was overall survival (OS) at 12 months (OS-12). Other outcomes were overall objective response rate (ORR), median progression-free survival (PFS), PFS at 6 months (PFS-6), and safety. Exploratory genomic and immune profiling using CITEseq was performed in 16 patients. Results: 90 patients were enrolled with median age of 60.6 years (range 27-86). 35 patients were MGMT methylated, 53 were unmethylated and 2 were indeterminate. Final analysis had 87 evaluable patients, with median follow-up of 146 days (range 7-1239). There were 3 complete responses (3.4%), 29 partial responses (33.3%), 37 stable disease (42.5%), and 18 progressive disease (20.7%). OS-12 was 41.1% and 37.7% in standard-dose and low-dose arms respectively, with PFS and ORR data in table. Post-hoc analysis demonstrated survival benefit for patients aged ≥60 years in standard-dose arm. Most frequent toxicities (>20%) included fatigue (45.6%), proteinuria (34.4 %), diarrhea (28.9%), hypertension (23.3%) and lipase increase (21.1%). Grade 3-4 toxicities included hypertension (7.8%), fatigue (5.6) and non-neurological toxicities. Differential changes across trial arms were seen in myeloid-derived suppressor cells (MDSCs) post-therapy. Network medicine identified VEGF as target to reduce MDSCs. VEGF was found expressed by MDSCs of mainly responder patients. Differential gene expression analysis identified increased pro-inflammatory gene signatures in standard-dose arm. Conclusions: Overall PFS and OS were similar for nivolumab plus standard- or low-dose bevacizumab for rGBM, with post-hoc survival benefit seen in standard-dose arm in elderly. Standard-dose bevacizumab was associated with increased systemic inflammatory response and reduced immunosuppressive MDSCs. Clinical trial information: NCT03452579 . [Table: see text]

Equine Endothelial Cells Show Pro-Angiogenic Behaviours in Response to Fibroblast Growth Factor 2 but Not Vascular Endothelial Growth Factor A.

Understanding the factors which control endothelial cell (EC) function and angiogenesis is crucial for developing the horse as a disease model, but equine ECs remain poorly studied. In this study, we have optimised methods for the isolation and culture of equine aortic endothelial cells (EAoECs) and characterised their angiogenic functions in vitro. Mechanical dissociation, followed by magnetic purification using an anti-VE-cadherin antibody, resulted in EC-enriched cultures suitable for further study. Fibroblast growth factor 2 (FGF2) increased the EAoEC proliferation rate and stimulated scratch wound closure and tube formation by EAoECs on the extracellular matrix. Pharmacological inhibitors of FGF receptor 1 (FGFR1) (SU5402) or mitogen-activated protein kinase (MEK) (PD184352) blocked FGF2-induced extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation and functional responses, suggesting that these are dependent on FGFR1/MEK-ERK signalling. In marked contrast, vascular endothelial growth factor-A (VEGF-A) had no effect on EAoEC proliferation, migration, or tubulogenesis and did not promote ERK1/2 phosphorylation, indicating a lack of sensitivity to this classical pro-angiogenic growth factor. Gene expression analysis showed that unlike human ECs, FGFR1 is expressed by EAoECs at a much higher level than both VEGF receptor (VEGFR)1 and VEGFR2. These results suggest a predominant role for FGF2 versus VEGF-A in controlling the angiogenic functions of equine ECs. Collectively, our novel data provide a sound basis for studying angiogenic processes in horses and lay the foundations for comparative studies of EC biology in horses versus humans.

Placental growth factor promotes neural invasion and predicts disease prognosis in resectable pancreatic cancer

BackgroundSurgery represents the only curative treatment option for pancreatic ductal adenocarcinoma (PDAC), but recurrence in more than 85% of patients limits the success of curative-intent tumor resection. Neural invasion (NI), particularly the spread of tumor cells along nerves into extratumoral regions of the pancreas, constitutes a well-recognized risk factor for recurrence. Hence, monitoring and therapeutic targeting of NI offer the potential to stratify recurrence risk and improve recurrence-free survival. Based on the evolutionary conserved dual function of axon and vessel guidance molecules, we hypothesize that the proangiogenic vessel guidance factor placental growth factor (PlGF) fosters NI. To test this hypothesis, we correlated PlGF with NI in PDAC patient samples and functionally assessed its role for the interaction of tumor cells with nerves.MethodsSerum levels of PlGF and its soluble receptor sFlt1, and expression of PlGF mRNA transcripts in tumor tissues were determined by ELISA or qPCR in a retrospective discovery and a prospective validation cohort. Free circulating PlGF was calculated from the ratio PlGF/sFlt1. Incidence and extent of NI were quantified based on histomorphometric measurements and separately assessed for intratumoral and extratumoral nerves. PlGF function on reciprocal chemoattraction and directed neurite outgrowth was evaluated in co-cultures of PDAC cells with primary dorsal-root-ganglia neurons or Schwann cells using blocking anti-PlGF antibodies.ResultsElevated circulating levels of free PlGF correlated with NI and shorter overall survival in patients with PDAC qualifying for curative-intent surgery. Furthermore, high tissue PlGF mRNA transcript levels in patients undergoing curative-intent surgery correlated with a higher incidence and greater extent of NI spreading to tumor-distant extratumoral nerves. In turn, more abundant extratumoral NI predicted shorter disease-free and overall survival. Experimentally, PlGF facilitated directional and dynamic changes in neurite outgrowth of primary dorsal-root-ganglia neurons upon exposure to PDAC derived guidance and growth factors and supported mutual chemoattraction of tumor cells with neurons and Schwann cells.ConclusionOur translational results highlight PlGF as an axon guidance factor, which fosters neurite outgrowth and attracts tumor cells towards nerves. Hence, PlGF represents a promising circulating biomarker of NI and potential therapeutic target to improve the clinical outcome for patients with resectable PDAC.Graphical

#2396 The potential of human-derived macrophages to study autosomal dominant polycystic kidney disease (ADPKD)

Abstract Background and Aims Macrophages are the most prevalent immune cell type in the renal parenchyma in ADPKD. Depletion of macrophages in animal models has shown a dramatic attenuation in cystic burden, suggesting a disease promoting effect of this cell type. So far, studies investigating interaction between epithelial cells and immune cells in the field of ADPKD were performed with mice-derived myeloid cells. We present a new model using human-derived immune cells and tubular epithelial cells for the study of ADPKD. Method We established a technique to isolate CD14+ monocytes from the peripheral blood of young ADPKD patients and healthy controls. Methodology was optimized to create monocyte-derived macrophagesin vitro. Mature macrophages were checked for surface marker expression (CD14, CD11b, CD206, CD163, HLA-DR, D80, and CD86) & cytokine-release after stimulation with LPS, IL-4 and LPS + Nigericin. Co-cultures were performed using human-derived monocytes in combination with double immortalized cystic cells (ciCC) or healthy-kidney derived cell lines (HKT). Differentiation and polarization of macrophages was checked using flowcytometry. Results We observed low expression of PKD1 and PKD2 transcripts in primary, human-derived monocytes of ADPKD patients and controls. The expression of PKD1 and PKD2 increased after differentiation into macrophages. There were no clear differences between ADPKD and control-derived macrophages in the differentiation capacity, cell survival nor surface marker expression before and after differentiation. After stimulation with LPS & LPS + Nigericin ADPKD-derived macrophages showed significantly lower expression of pro-inflammatory cytokines (MIP-1α, IP-10, IL-23 and TNFα) as compared to controls. There was no difference in anti-inflammatory cytokine release. Healthy primary monocytes cultured in direct contact with conditionally immortalized cystic cells (ciCC) had a very good survival and showed differentiation into macrophages with high expression of M2-markers (CD163 and CD206) and the activation marker (HLA-DR). A similar differentiation pattern was seen when monocytes were cultured in the (cell-free) conditioned medium of the ciCC harvested after 72 h, pointing to a soluble factor produced by the cystic cells that induces monocyte differentiation and macrophage polarization. In accordance, unbiased multiplex cytokine analysis (Target 48-panel) of supernatants of ciCC, showed higher levels of macrophage colony-stimulating factor (M-CSF/CSF-1), TNF-α, hepatocyte growth factor (HGF) and MMP-1 as compared to those of HKT. Primary monocytes cultured with the conditioned medium harvested after 72 h from ciCCs (n=4) showed better overall survival than those cultured with medium from and HKT (n=4). In addition, a tendency towards higher expression of the classical M2-markers (CD163, CD206) and the activation marker (HLA-DR) in ciCCs versus HKT was seen. Conclusion There is wealth of data supporting the indispensable role of myeloid cells in disease progression of ADPKD. We established a unique in vitro fully human co-culture model for ADPKD. Cystic cells produce pro-proliferative and pro-angiogenic factors, thus actively shaping their microenvironment and creating the ideal circumstances for infiltrating monocytes/macrophages to proliferate and to destruct the kidney. This mechanism is very similar to what is seen in the cancer microenvironment. Further experiments will help to elucidate the complex interplay between different cell-types in the microenvironment in ADPKD kidneys.

Endothelial PDGF-D contributes to neurovascular protection after ischemic stroke by rescuing pericyte functions

Ischemic stroke induces neovascularization of the injured tissue as an attempt to promote structural repair and neurological recovery. Angiogenesis is regulated by pericytes that potently react to ischemic stroke stressors, ranging from death to dysfunction. Platelet-derived growth factor (PDGF) receptor (PDGFR)β controls pericyte survival, migration, and interaction with brain endothelial cells. PDGF-D a specific ligand of PDGFRβ is expressed in the brain, yet its regulation and role in ischemic stroke pathobiology remains unexplored. Using experimental ischemic stroke mouse model, we found that PDGF-D is transiently induced in brain endothelial cells at the injury site in the subacute phase. To investigate the biological significance of PDGF-D post-ischemic stroke regulation, its subacute expression was either downregulated using siRNA or upregulated using an active recombinant form. Attenuation of PDGF-D subacute induction exacerbates neuronal loss, impairs microvascular density, alters vascular permeability, and increases microvascular stalling. Increasing PDGF-D subacute bioavailability rescues neuronal survival and improves neurological recovery. PDGF-D subacute enhanced bioavailability promotes stable neovascularization of the injured tissue and improves brain perfusion. Notably, PDGF-D enhanced bioavailability improves pericyte association with brain endothelial cells. Cell-based assays using human brain pericyte and brain endothelial cells exposed to ischemia-like conditions were applied to investigate the underlying mechanisms. PDGF-D stimulation attenuates pericyte loss and fibrotic transition, while increasing the secretion of pro-angiogenic and vascular protective factors. Moreover, PDGF-D stimulates pericyte migration required for optimal endothelial coverage and promotes angiogenesis. Our study unravels new insights into PDGF-D contribution to neurovascular protection after ischemic stroke by rescuing the functions of pericytes.

Angiogenesis Still Plays a Crucial Role in Human Melanoma Progression.

Angiogenesis plays a pivotal role in tumor progression, particularly in melanoma, the deadliest form of skin cancer. This review synthesizes current knowledge on the intricate interplay between angiogenesis and tumor microenvironment (TME) in melanoma progression. Pro-angiogenic factors, including VEGF, PlGF, FGF-2, IL-8, Ang, TGF-β, PDGF, integrins, MMPs, and PAF, modulate angiogenesis and contribute to melanoma metastasis. Additionally, cells within the TME, such as cancer-associated fibroblasts, mast cells, and melanoma-associated macrophages, influence tumor angiogenesis and progression. Anti-angiogenic therapies, while showing promise, face challenges such as drug resistance and tumor-induced activation of alternative angiogenic pathways. Rational combinations of anti-angiogenic agents and immunotherapies are being explored to overcome resistance. Biomarker identification for treatment response remains crucial for personalized therapies. This review highlights the complexity of angiogenesis in melanoma and underscores the need for innovative therapeutic approaches tailored to the dynamic TME.

Preventive effects of vasohibin-2-targeting peptide vaccine for diabetic nephropathy.

Diabetic nephropathy remains the leading cause of end-stage kidney disease in many countries, and additional therapeutic targets are needed to prevent its development and progression. Some angiogenic factors are involved in the pathogenesis of diabetic nephropathy. Vasohibin-2 (VASH2) is a novel proangiogenic factor, and our previous study showed that glomerular damage is inhibited in diabetic Vash2 homozygous knockout mice. Therefore, we established a VASH2-targeting peptide vaccine as a tool for anti-VASH2 therapy in diabetic nephropathy. In this study, the preventive effects of the VASH2-targeting peptide vaccine against glomerular injury were examined in a streptozotocin (STZ)-induced diabetic mouse model. The mice were subcutaneously injected with the vaccine at two doses 2 wk apart and then intraperitoneally injected with 50 mg/kg STZ for 5 consecutive days. Glomerular injury was evaluated 20 wk after the first vaccination. Treatment with the VASH2-targeting peptide vaccine successfully induced circulating anti-VASH2 antibody without inflammation in major organs. Although the vaccination did not affect blood glucose levels, it significantly prevented hyperglycemia-induced increases in urinary albumin excretion and glomerular volume. The vaccination did not affect increased VASH2 expression but significantly inhibited renal angiopoietin-2 (Angpt2) expression in the diabetic mice. Furthermore, it significantly prevented glomerular macrophage infiltration. The preventive effects of vaccination on glomerular injury were also confirmed in db/db mice. Taken together, the results of this study suggest that the VASH2-targeting peptide vaccine may prevent diabetic glomerular injury in mice by inhibiting Angpt2-mediated microinflammation.NEW & NOTEWORTHY This study demonstrated preventive effects of VASH2-targeting peptide vaccine therapy on albuminuria and glomerular microinflammation in STZ-induced diabetic mouse model by inhibiting renal Angpt2 expression. The vaccination was also effective in db/db mice. The results highlight the importance of VASH2 in the pathogenesis of early-stage diabetic nephropathy and the practicability of anti-VASH2 strategy as a vaccine therapy.

Role of soluble fms-like tyrosine kinase-1/placental growth factor ratio along with uterine artery Doppler for the prediction of pre-eclampsia – A case–control study

Role of soluble fms-like tyrosine kinase-1/placental growth factor ratio along with uterine artery Doppler for the prediction of pre-eclampsia – A case–control study

Abstract 2037: Investigating Changes In Satellite Cell Proliferation And Angiogenic Potential In Response To Exercise

Background: Peripheral Artery Disease (PAD) is an atherosclerotic process leading to vessel obstruction. Supervised walking exercise is a first-line treatment to improve walking ability and quality of life scores of patients. Despite its known benefits, the underlying mechanism is not fully understood. Thus, this study focuses on satellite cells (SCs), known for their role in muscular regeneration but also found to have an under investigated role as mediators of vascular growth through the secretion of cytokines and growth factors. Previous in vivo models have demonstrated increased migration of endothelial cells and smooth muscle cells, as well as blood vessel growth in response to SC activity. Aims: The project aims to enhance the efficacy of exercise therapy through a better understanding satellite cell biology and their pro-vasculogenic ability. Hypothesis: Exercise therapy will enhance SCs proliferation and secretion of paracrine growth factors inducing increased cell migration and vascular growth. Methods: Aortic Ring Assay: Murine thoracic aortic sections were embedded in a Type I collagen matrix in 96-wells. The wells were seeded with Opti-MEM (control), Opti-MEM+0.3% Vascular Endothelial Growth Factors (VEGF)(+ control), SC Conditioned Media (CM) (24hr incubation, 2M cells/plate), or denatured CM (56°C, 1 hr). The total length and number of branches were measured via ImageJ. Flow Cytometry: 18+ months old C57BL/6 were housed with voluntary running wheels with activity counter until day 28. Satellite cells were mechanically and chemically digested and were labeled as: CD11b-, CD45-, CD31-, Sca1-, and α7-integrin+ at 7-day intervals. Results: Aortic Ring Assay: SC CM showed significantly higher total branch length (13381.89±2470.44μm, n=6) (****p<0.0001) and numbers (19.24±3.87, n=6) (**p<0.01) compared to control (2158.18±382.31μm, n=9), (5.76±0.81, n=9). Flow Cytometry: SC proportion on day 21 (2.55±0.14,n=2) was significantly higher than control (1.54±0.298,n=8)(*p<0.05). Conclusion(s): These findings indicate that SC CM induces angiogenesis by secreting pro-angiogenic factors. The angiogenic activities could potentially be heightened through the proliferation of SCs under exercise therapy.

FTO elicits tumor neovascularization in cancer-associated fibroblasts through eliminating m6A modifications of multiple pro-angiogenic factors

Cancer-associated fibroblasts (CAFs) exhibit notable versatility, plasticity, and robustness, actively participating in cancer progression through intricate interactions within the tumor microenvironment (TME). N6-methyladenosine (m6A) modification is the most prevalent modification in eukaryotic mRNA, playing essential roles in mRNA metabolism and various biological processes. Howbeit, the precise involvement of m6A in CAF activation remains enigmatic. In this study, we revealed that the m6A demethylase FTO supports CAF-mediated angiogenesis through activation of EGR1 and VEGFA in conjunctival melanoma (CoM). First, single-cell transcriptome analysis revealed that FTO was specifically upregulated in the CAF population, thereby contributing to the hypo-m6A status in the TME of CoM. Moreover, CAFs of CoM displayed extensive proangiogenic potential, which was largely compromised by FTO inhibition, both in vitro and in vivo. By employing multi-omics analysis, we showed that FTO effectively eliminates the m6A modifications of VEGFA and EGR1. This process subsequently disrupts the YTHDF2-dependent mRNA decay pathway, resulting in increased mRNA stability and upregulated expression of these molecules. Collectively, our findings initially indicate that the upregulation of FTO plays a pivotal role in tumor development by promoting CAF-mediated angiogenesis. Therapeutically, targeting FTO may show promise as a potential antiangiogenic strategy to optimize cancer treatment.

Immunohistochemical evaluation of tumor hypoxia and angiogenesis: Pathological significance and prognostic role in head and neck squamous cell carcinomas

Objectives: Tumor hypoxia and angiogenesis have been implicated in therapeutic resistance of head and neck squamous cell carcinomas (HNSCCs). Immunohistochemical evaluation of hypoxia-inducible factor-1 alpha (HIF-1 α), a hypoxia transcription factor, and vascular endothelial growth factor (VEGF), a hypoxia-responsive pro-angiogenic factor, can be exploited for prognostication and guiding treatment intensification or de-escalation decisions in HNSCC patients. The purpose of the study is to evaluate the immunohistochemical expression patterns of HIF-1 α and VEGF and the microvessel density (MVD) for angiogenesis in HNSCC and assess their pathological significance and prognostic role. Materials and Methods: In this cross-sectional study, immunohistochemical expression of HIF-1 α, VEGF, and MVD through Cluster of Differentiation (CD31) was evaluated in paraffin-embedded tumor resection tissue of 44 patients with HNSCC. Associations among HIF-1 α, VEGF, and MVD with clinicopathological variables were assessed. Statistical Analysis: For assessment of association between HIF-1α, VEGF and MVD by CD 31 immunohistochemical markers and other clinicopathological variables Pearson’s chi-square test and Fisher’s exact tests were used. Analysis of survival was done using Kaplan-Meier statistics. Also, the univariate and multivariate analysis were performed using the Cox proportional hazard regression model for the calculation of hazard ratios. Results: Nuclear expression of HIF-1 α showed significant association with MVD (P = 0.007) and cytoplasmic expression of HIF-1 α with histologic grade (P = 0.03). Overexpression of HIF-1 α was more frequent in T3/T4 stage. In addition to cytoplasmic staining, VEGF showed a unique nuclear expression pattern in four cases of advanced disease with nodal metastasis. Logistic regression analysis showed tumors with nuclear overexpression of HIF-1 α to have increased MVD (P = 0.05), and tumors with higher MVD to have a presence of lymphovascular invasion (P = 0.014). Multivariate analysis showed HIF-1 α nuclear overexpression to be significantly associated with decreased survival of patients (P = 0.05). Conclusions: Immunohistochemical overexpression of HIF-1 α and MVD quantification can serve as cost-effective tools for prognostication and treatment modification of HNSCC patients in resourcelimited settings.

Metformin inhibits activation of NLRP3 inflammasome and inflammatory response in preeclamptic rats

Backgroud: It is widely acknowledged that Metformin (MET), an established medication for managing type 2 diabetes, possesses diverse pharmacological effects. This study aims to investigate the protective effects of MET against Nω-Nitro-L-arginine methyl ester (L-NAME)-induced preeclampsia (PE). MethodsSprague Dawley (SD) rats were exposed to 200 mg/kg L-NAME with or without prior MET treatment. Histopathological analysis was performed using Hematoxylin and Eosin staining. Serum levels of inflammatory, antiangiogenic, and angiogenic factors were quantified using ELISA kits. Immunohistochemistry (IHC) staining was employed to observe NLRP3 and IL-1β expressions in placental tissues. Western blot and Quantitative Real-Time PCR (q-PCR) analyses were conducted to assess protein and mRNA expressions of NLRP3, caspase-1, ASC, and IL-1β. ResultsWe found that MET could mitigate placental histopathological deterioration and improve pregnancy outcomes in L-NAME-induced PE rat models. MET not only suppressed L-NAME-induced elevation of antiangiogenic factors but also stimulated the production of pro-angiogenic factors. Additionally, MET treatment reversed the excessive inflammatory response induced by L-NAME. Furthermore, MET inhibited the activation of the NLRP3 inflammasome triggered by L-NAME, evidenced by the downregulation of NLRP3 expression, caspase-1, and IL-1β. ConclusionsMET demonstrates a protective effect against L-NAME-induced PE rats, potentially mediated through inhibition of the inflammatory response, downregulation of NLRP3 inflammasome expression in the placenta, and regulation of the balance between anti-angiogenic and pro-angiogenic factors.

Angiopoietin 1 Relieves Osteolysis by Promoting Macrophage Mitophagy Through the TBK1-SQSTM1 Pathway to Inhibit AIM2 Inflammasome-Mediated Pyroptosis.

Osteolysis resulting from wear particles and subsequent aseptic loosening is a leading cause of revision surgery of artificial joints. The underlying pathogenesis of particle-induced osteolysis (PPO) has remained largely uncertain. Addressing how to mitigate osteolysis caused by wear particles presents a significant challenge for orthopedic surgeons. This study aimed to explore the molecular mechanism by which Angiopoietin (Ang-1) inhibits osteoclast activation to alleviate osteolysis. RAW264.7 mouse macrophages were stimulated with LPS or RANKL to induce osteoclast formation. Additionally, titanium (Ti) particles (50mg) were subperiosteally implanted around the cranial suture of mice to establish a calvarial osteolysis model. Ang-1, a member of the pro-angiogenic factor protein family and an important inflammatory regulator molecule, was utilized in this model. TRAP staining was utilized to detect osteoclast activation, while a western blot was conducted to identify key proteins associated with mitophagy and pyroptosis. Scanning electron microscopy was employed to observe the morphology and dimensions of Ti particles. Additionally, a combination of micro-CT, H&E, Masson's trichrome, and immunohistochemical staining techniques were applied to analyze the calvarial samples. Results indicated that Ang-1 could inhibit LPS- or RANKL-induced osteoclastogenesis and alleviate Ti particle-induced calvarial osteolysis in mice. TBK-1, a key signaling molecule involved in initiating mitophagy, was found to be mechanistically enhanced by Ang-1 through promoting TBK-1 phosphorylation in macrophages. This process inhibited AIM2 inflammasome-mediated pyroptosis and impeded osteoclastogenesis. Overall, this research uncovers a novel mechanism by which Ang-1 can attenuate inflammatory osteolysis, potentially offering a new therapeutic approach for PPO.

Signaling crosstalk between tumor endothelial cells and immune cells in the microenvironment of solid tumors.

Tumor-associated endothelial cells (TECs) are crucial mediators of immune surveillance and immune escape in the tumor microenvironment (TME). TECs driven by angiogenic growth factors form an abnormal vasculature which deploys molecular machinery to selectively promote the function and recruitment of immunosuppressive cells while simultaneously blocking the entry and function of anti-tumor immune cells. TECs also utilize a similar set of signaling regulators to promote the metastasis of tumor cells. Meanwhile, the tumor-infiltrating immune cells further induce the TEC anergy by secreting pro-angiogenic factors and prevents further immune cell penetration into the TME. Understanding the complex interactions between TECs and immune cells will be needed to successfully treat cancer patients with combined therapy to achieve vasculature normalization while augmenting antitumor immunity. In this review, we will discuss what is known about the signaling crosstalk between TECs and tumor-infiltrating immune cells to reveal insights and strategies for therapeutic targeting.

Enhanced Vascular-like Network Formation of Encapsulated HUVECs and ADSCs Coculture in Growth Factors Conjugated GelMA Hydrogels.

Tissue engineering primarily aimed to alleviate the insufficiency of organ donations worldwide. Nonetheless, the survival of the engineered tissue is often compromised due to the complexity of the natural organ architectures, especially the vascular system inside the organ, which allows food-waste transfer. Thus, vascularization within the engineered tissue is of paramount importance. A critical aspect of this endeavor is the ability to replicate the intricacies of the extracellular matrix and promote the formation of functional vascular networks within engineered constructs. In this study, human adipose-derived stem cells (hADSCs) and human umbilical vein endothelial cells (HUVECs) were cocultured in different types of gelatin methacrylate (GelMA). In brief, pro-angiogenic signaling growth factors (GFs), vascular endothelial growth factor (VEGF165) and basic fibroblast growth factor (bFGF), were conjugated onto GelMA via an EDC/NHS coupling reaction. The GelMA hydrogels conjugated with VEGF165 (GelMA@VEGF165) and bFGF (GelMA@bFGF) showed marginal changes in the chemical and physical characteristics of the GelMA hydrogels. Moreover, the conjugation of these growth factors demonstrated improved cell viability and cell proliferation within the hydrogel construct. Additionally, vascular-like network formation was observed predominantly on GelMA@GrowthFactor (GelMA@GF) hydrogels, particularly on GelMA@bFGF. This study suggests that growth factor-conjugated GelMA hydrogels would be a promising biomaterial for 3D vascular tissue engineering.

Improving the mechanical and biological functions of cell sheet constructs: The interplay of human-derived periodontal ligament stem cells, endothelial cells and plasma rich in growth factors

ObjectiveThe aim of this study was to produce and characterize triple-layered cell sheet constructs with varying cell compositions combined or not with the fibrin membrane scaffold obtained by the technology of Plasma Rich in Growth Factors (mPRGF). Materials and methodsHuman primary cultures of periodontal ligament stem cells (hPDLSCs) were isolated, and their stemness nature was evaluated. Three types of triple-layered composite constructs were generated, composed solely of hPDLSCs or combined with human umbilical vein endothelial cells (HUVECs), either as a sandwiched endothelial layer or as coculture sheets of both cell phenotypes. These three triple-layered constructs were also manufactured using mPRGF as cell sheets’ support. Necrosis, glucose consumption, secretion of extracellular matrix proteins and synthesis of proangiogenic factors were determined. Histological evaluations and proteomic analyses were also performed. ResultsThe inclusion of HUVECs did not clearly improve the properties of the multilayered constructs and yet hindered their optimal conformation. The presence of mPRGF prevented the shrinkage of cell sheets, stimulated the metabolic activity and increased the matrix synthesis. At the proteome level, mPRGF conferred a dramatic advantage to the hPDLSC constructs in their ability to provide a suitable environment for tissue regeneration by inducing the expression of proteins necessary for bone morphogenesis and cellular proliferation. ConclusionshPDLSCs’ triple-layer construct onto mPRGF emerges as the optimal structure for its use in regenerative therapeutics. Clinical relevanceThese results suggest the suitability of mPRGF as a promising tool to support cell sheet formation by improving their handling and biological functions.

DLC1 promotes mechanotransductive feedback for YAP via RhoGAP-mediated focal adhesion turnover.

Angiogenesis is a tightly controlled dynamic process demanding a delicate equilibrium between pro-angiogenic signals and factors that promote vascular stability. The spatiotemporal activation of the transcriptional co-factors YAP (herein referring to YAP1) and TAZ (also known WWTR1), collectively denoted YAP/TAZ, is crucial to allow for efficient collective endothelial migration in angiogenesis. The focal adhesion protein deleted-in-liver-cancer-1 (DLC1) was recently described as a transcriptional downstream target of YAP/TAZ in endothelial cells. In this study, we uncover a negative feedback loop between DLC1 expression and YAP activity during collective migration and sprouting angiogenesis. In particular, our study demonstrates that signaling via the RhoGAP domain of DLC1 reduces nuclear localization of YAP and its transcriptional activity. Moreover, the RhoGAP activity of DLC1 is essential for YAP-mediated cellular processes, including the regulation of focal adhesion turnover, traction forces, and sprouting angiogenesis. We show that DLC1 restricts intracellular cytoskeletal tension by inhibiting Rho signaling at the basal adhesion plane, consequently reducing nuclear YAP localization. Collectively, these findings underscore the significance of DLC1 expression levels and its function in mitigating intracellular tension as a pivotal mechanotransductive feedback mechanism that finely tunes YAP activity throughout the process of sprouting angiogenesis.

Targeted delivery of factors and cells for improving cardiac tissue regeneration and heart function following myocardial infarction

Following myocardial infarction (MI), cardiac tissue undergoes irreversible cellular alterations, with cardiomyocytes being replaced by fibrotic tissue. In order to improve tissue regeneration, a previously characterized chitosan-based cryogel, which was designed by our group, was used. The treatment regimen involved the sequential delivery of the cryogel loaded with specific cytokines and growth factors, followed by a separate injection of pre-differentiated cells. Initially, the cryogel loaded with interleukin-10 and transforming growth factor-β was injected into infarcted tissue immediately after MI induction, targeting the acute inflammatory response. On day four post-MI, a second injection was administered, this time utilizing cryogel loaded with vascular endothelial growth factor and fibroblast growth factor-2, aimed at promoting tissue regeneration and angiogenesis. Subsequently, on day six post-MI, the experimental group received cardiomyocyte-like cells, smooth muscle cells, and endothelial cells. The purpose of these cells, in synergy with cytokines and growth factors, was to repopulate the lost cellular populations, thereby enhancing myocardial repair. The treatment improved myocardial tissue regeneration, increased cardiac output, ejection fraction, and reduced fibrotic regions. Thus, the chitosan-based cryogel, enriched with anti-inflammatory and proangiogenic factors and supplemented with pre-differentiated cells, offers a promising platform for controlled release of therapeutics, promoting substantial tissue repair and regeneration following MI.

Decoding Tumor Angiogenesis for Therapeutic Advancements: Mechanistic Insights.

Tumor angiogenesis, the formation of new blood vessels within the tumor microenvironment, is considered a hallmark of cancer progression and represents a crucial target for therapeutic intervention. The tumor microenvironment is characterized by a complex interplay between proangiogenic and antiangiogenic factors, regulating the vascularization necessary for tumor growth and metastasis. The study of angiogenesis involves a spectrum of techniques, spanning from biomarker assessment to advanced imaging modalities. This comprehensive review aims to provide insights into the molecular intricacies, regulatory dynamics, and clinical implications of tumor angiogenesis. By delving into these aspects, we gain a deeper understanding of the processes driving vascularization in tumors, paving the way for the development of novel and effective antiangiogenic therapies in the fight against cancer.

An injectable anti-vascularization functionalized hydrogel for degenerative nucleus pulposus repair

Neovascularization and inflammatory cell invasion within the nucleus pulposus (NP) constitute pivotal pathological changes during the acceleration stage of intervertebral disc degeneration (IDD). Mesenchymal stem cells (MSCs), renowned for their remarkable capacity in intervertebral disc (IVD) regeneration, also exhibit the capability to secrete pro-angiogenic factors, expediting IDD progression under hypoxic conditions. Consequently, we developed a hydrogel comprised of methacrylated hyaluronic acid (HAMA), rat tail collagen I (COL), and MSCs, incorporating the vascular endothelial growth factor receptor (VEGFR) inhibitor cabozantinib (Cabo@HAMA-COL/MSCs hydrogel). This innovative construct aimed to facilitate NP regeneration while mitigating vascularization and inflammation. Our findings revealed that the hydrogel aptly mimicked the mechanical characteristics of NP tissue, exhibiting injectability, low cytotoxicity, and the preservation of the cellular phenotype of NP cells. Co-culturing of MSCs and human umbilical vein endothelial cells (HUVECs) promoted migration, tube formation, and sprouting of HUVECs, which will be inhibited by cabozantinib. In vivo experiments demonstrated that Cabo@HAMA-COL/MSCs hydrogel maintained disc height, protected NP, and alleviated vascularization and inflammation in a puncture-induced rat caudal IDD model. Consequently, our results substantiate that Cabo@HAMA-COL/MSCs hydrogel can prevent IDD degeneration by ameliorating the vascularization-inflammation pathological microenvironment, offering a promising therapeutic strategy for IDD.