Role of regulatory T cells in mouse lung development.

Regulatory T cells (Tregs) constitute a specialized subset of T cells with dual immunoregulatory and modulatory functions. Recent studies have reported that Tregs mediate immune responses and regulate the development and repair processes in non-lymphoid tissues, including bone and cardiac muscle. Additionally, Tregs facilitate the repair and regeneration of damaged lung tissues. However, limited studies have examined the role of Tregs in pulmonary development. This study aimed to evaluate the role of Tregs in pulmonary development by investigating the dynamic alterations in Tregs and their hallmark cellular factor Forkhead box P3 (Foxp3) at various stages of murine lung development and establishing a murine model of anti-CD25 antibody-induced Treg depletion. During the early stages of murine lung development, especially the canalicular and saccular stages, the levels of Treg abundance and expression of Foxp3 and transforming growth factor-ß (TGF-ß) were upregulated. This coincided with the proliferation period of alveolar epithelial cells and vascular endothelial cells, indicating an adaptation to the dynamic lung developmental processes. Furthermore, the depletion of Tregs disrupted lung tissue morphology and downregulated lung development-related factors, such as surfactant protein C (SFTPC), vascular endothelial growth factor A (VEGFA) and platelet endothelial cell adhesion molecule-1 (PECAM1/CD31). These findings suggest that Tregs promote murine lung development.

Doxazosin inhibits vasculogenic mimicry in human non­small cell lung cancer through inhibition of the VEGF­A/VE­cadherin/mTOR/MMP pathway.

Lung cancer is the leading cause of cancer-related death worldwide, and ~85% of lung cancers are non-small cell lung cancer (NSCLC), which has a low 5-year overall survival rate and high mortality. Several therapeutic strategies have been developed, such as targeted therapy, immuno-oncotherapy and combination therapy. However, the low survival rate indicates the urgent need for new NSCLC treatments. Vasculogenic mimicry (VM) is an endothelial cell-free tumor blood supply system of aggressive and metastatic tumor cells present during tumor neovascularization. VM is clinically responsible for tumor metastasis and resistance, and is correlated with poor prognosis in NSCLC, making it a potential therapeutic target. In the present study, A549 cells formed glycoprotein-rich lined tubular structures, and transcript levels of VM-related genes were markedly upregulated in VM-forming cells. Based on a drug repurposing strategy, it was demonstrated that doxazosin (an antihypertensive drug) displayed inhibitory activity on VM formation at non-cytotoxic concentrations. Doxazosin significantly reduced the levels of vascular endothelial growth factor A (VEGF-A) and matrix metalloproteinase-2 (MMP-2) in the cell media during VM formation. Further experiments revealed that the protein expression levels of VEGF-A and vascular endothelial-cadherin (VE-cadherin), which contribute to tumor aggressiveness and VM formation, were downregulated following doxazosin treatment. Moreover, the downstream signaling Ephrin type-A receptor 2 (EphA2)/AKT/mTOR/MMP/Laminin-5γ2 network was inhibited in response to doxazosin treatment. In conclusion, the present study demonstrated that doxazosin displayed anti-VM activity in an NSCLC cell model through the downregulation of VEGF-A and VE-cadherin levels, and the suppression of signaling pathways related to the receptor tyrosine kinase, EphA2, protein kinases, AKT and mTOR, and proteases, MMP-2 and MMP-9. These results support the add-on anti-VM effect of doxazosin as a potential agent against NSCLC.

Ubiquitin ligase MDM2 mediates endothelial inflammation in Kawasaki disease vasculitis development.

Background: Kawasaki disease (KD) often complicates coronary artery lesions (CALs). Despite the established significance of STAT3 signaling during the acute phase of KD and signal transducer and activator of transcription 3 (STAT3) signaling being closely related to CALs, it remains unknown whether and how STAT3 was regulated by ubiquitination during KD pathogenesis. Methods: Bioinformatics and immunoprecipitation assays were conducted, and an E3 ligase, murine double minute 2 (MDM2) was identified as the ubiquitin ligase of STAT3. The blood samples from KD patients before and after intravenous immunoglobulin (IVIG) treatment were utilized to analyze the expression level of MDM2. Human coronary artery endothelial cells (HCAECs) and a mouse model were used to study the mechanisms of MDM2-STAT3 signaling during KD pathogenesis. Results: The MDM2 expression level decreased while the STAT3 level and vascular endothelial growth factor A (VEGFA) level increased in KD patients with CALs and the KD mouse model. Mechanistically, MDM2 colocalized with STAT3 in HCAECs and the coronary vessels of the KD mouse model. Knocking down MDM2 caused an increased level of STAT3 protein in HCAECs, whereas MDM2 overexpression upregulated the ubiquitination level of STAT3 protein, hence leading to significantly decreased turnover of STAT3 and VEGFA. Conclusions: MDM2 functions as a negative regulator of STAT3 signaling by promoting its ubiquitination during KD pathogenesis, thus providing a potential intervention target for KD therapy.

Perifoveal retinal thickness changes after intravitreal aflibercept injection for choroidal neovascularization in age-related macular degeneration.

OBJECTIVE: There has been some concern that anti-vascular growth factor treatment accelerates the development of macular atrophy in eyes with neovascular age-related macular degeneration. During the treatment with aflibercept, the thickness of choroid may decrease. This may lead to photoreceptor death. The rod cells are more susceptible to atrophic changes than cones during the disease. We aimed to find any thickness changes in the perifoveal outer nuclear layer, where the highest density of rods is found, during the aflibercept intravitreal injection therapy. MATERIALS AND METHODS: Retrospectively, forty-two patients who were treated for age-related macular degeneration with choroidal neovascularization were included in the study. After the first three loading doses, intravitreal injections were repeated every two months. Outer nuclear layer thicknesses were measured 2000 µm away from the center of the fovea with OCT, at a total of 20 points, located at 180 and 90°. The mean of these measurements was obtained before the treatment and 1 year after the therapy. Results were compared by using the Wilcoxon Rank Test. RESULTS: The mean visual acuity was 1,11±0,287 logMAR at the beginning and increased to 0,53±0,32 LogMAR after. Perifoveal thickness was significantly reduced when compared with the thickness before the treatment (p = 0.039, p < 0.05). This result was also significantly lower than the control group thicknesses (p = 0.035, p < 0.05). CONCLUSION: Anti-VEGFs can cause loss of phagocytic functions of RPE. The mechanism of the observed thinning of the ONL may be described as follows: VEGF emitted by the RPE normally helps to maintain the choriocapillaris. Thus, injecting an anti-VEGF intravitreally causes RPE atrophy, which leads to a decrease in the choroidal vascular index. This in turn causes first the rods, and later the cones, that are part of the outer nuclear layer, to start to die and disappear, hence the thinning of this layer. As aflibercept consists of parts of the extracellular domain of both the VEGFR1 and VEGFR2 receptors, that are held together by a human IgG1 backbone, this makes the binding of aflibercept to VEGF-A and VEGF-B stronger as compared to the binding of the previously used ranibizumab or bevacizumab (by nearly a factor 100 in the case of the most abundant isoform VEGF-A 165). Besides, aflibercept also binds very well to placental growth factor (PIGF), which is also associated with several ocular diseases.

[Retracted] Downregulation of microRNA-206 suppresses clear cell renal carcinoma proliferation and invasion by targeting vascular endothelial growth factor A.

Following the publication of the above paper, it was drawn to the Editor's attention by a concerned reader that there appeared to be several instances of overlapping data panels comparing between the Transwell invasion and migration assay images shown in Figs. 2E and 4G, such that data which were intended to show the results from differently performed experiments were apparently derived from a (much) smaller number of original sources. Given the number of cases of overlapping data panels both within and between this pair of figures in the article itself, the Editor of Oncology Reports has decided that this paper should be retracted from the Journal on the basis of a lack of confidence in the presented data. The authors were asked for an explanation to account for these concerns, but the Editorial Office did not receive a reply. The Editor apologizes to the readership for any inconvenience caused. [Oncology Reports 35: 1778-1786, 2016; DOI: 10.3892/or.2015.4538].

[Tetramethylpyrazine regulates angiogenesis of endothelial cells in cerebral ischemic stroke injury via SIRT1/VEGFA signaling pathway].

This study aims to investigate whether tetramethylpyrazine(TMP) can stimulate angiogenesis in cerebral microvascular endothelial cells and alleviate cerebral ischemic stroke(CIS) and to explore the underlying mechanisms. In the animal study, adult Sprague-Dawley rats(n=15) were assigned into sham surgery(sham), middle cerebral artery occlusion/reperfusion(MCAO/R), and MCAO/R+TMP(intraperitoneal injection of 20 mg·kg~(-1)) groups. The neurological function was evaluated by the Z-Longa method. The cerebral infarction volume was detected by TTC staining. Enzyme-linked immunosorbent assay(ELISA) was employed to detect the expression of vascular endothelial growth factor(VEGF), angiopoietin(Ang), and platelet-derived growth factor(PDGF). Immunofluorescence staining was employed to detect Ki67 and the expression of vascular endothelial growth factor A(VEGFA) and slient information regulator 1(SIRT1). Western blot was employed to determine the expression levels of VEGFA, SIRT1, angiopoietin-2(Ang-2), and platelet-derived growth factor B(PDGFB). In the cell study, mouse brain-derived endothelial cells(Bend.3) were cultured, and the optimal concentration of TMP was determined. Then, VEGF, Ang, and PDGF were detected by ELISA after the addition of cabozantinib. Western blot was employed to measure the expression of VEGFA, Ang-2, and PDGFB. Immunofluorescence staining was used to detect CD31, CD34, and Ki67, and the proliferation, migration, and tube formation ability of Bend.3 cells were observed in vitro. Western blot and immunofluorescence staining were performed to measure the expression of SIRT1 and VEGFA after addition of the SIRT1-specific inhibitor selisistat(EX-527). The results showed that compared with the sham group, the MCAO/R group had severe neurological function damage, increased infarction volume, up-regulated expression of VEGF, VEGFA, Ang, Ang-2, PDGF, and PDGFB, and down-regulated expression of Ki67 and SIRT1(P<0.01). Compared with the MCAO/R group, the MCAO/R+TMP group presented alleviated neurological function damage, reduced infarction volume, and activated expression of VEGF, VEGFA, Ang, Ang-2, PDGF, PDGFB, Ki67, and SIRT1(P<0.01). The cell experiments showed that compared with the normal group, Bend.3 cells were activated by oxygen glucose deprivation/reoxygenation(OGD/R) treatment(P<0.05, P<0.01). Compared with the OGD/R group, the OGD/R+TMP group upregulated the expression levels of VEGF, VEGFA, Ang, Ang-2, PDGF, PDGFB, SIRT1, Ki67, CD31, and CD34, enhanced the angiogenic ability of Bend.3 cells without being inhibited by BMS or EX-527(P<0.05, P<0.01, P<0.001). The results suggest that TMP can activate the SIRT1/VEGFA signaling pathway to stimulate angiogenesis and alleviate CIS injury.

Downregulation of VEGFA accelerates AGEs-mediated nucleus pulposus degeneration through inhibiting protective mitophagy in high glucose environments.

Intervertebral disc degeneration (IVDD) contributes largely to low back pain. Recent studies have highlighted the exacerbating role of diabetes mellitus (DM) in IVDD, mainly due to the influence of hyperglycemia (HG) or the accumulation of advanced glycation end products (AGEs). Vascular endothelial growth factor A (VEGFA) newly assumed a distinct impact in nonvascular tissues through mitophagy regulation. However, the combined actions of HG and AGEs on IVDD and the involved role of VEGFA remain unclear. We confirmed the potential relation between VEGFA and DM through bioinformatics and biological specimen detection. Then we observed that AGEs induced nucleus pulposus (NP) cell degeneration by upregulating cellular reactive oxygen species (ROS), and HG further aggravated ROS level through breaking AGEs-induced protective mitophagy. Furthermore, this adverse effect could be strengthened by VEGFA knockdown. Importantly, we identified that the regulation of VEGFA and mitophagy were vital mechanisms in AGEs-HG-induced NP cell degeneration through Parkin/Akt/mTOR and AMPK/mTOR pathway. Additionally, VEGFA overexpression through local injection with lentivirus carrying VEGFA plasmids significantly alleviated NP degeneration and IVDD in STZ-induced diabetes and puncture rat models. In conclusion, the findings first confirmed that VEGFA protects against AGEs-HG-induced IVDD, which may represent a therapeutic strategy for DM-related IVDD.

TENAYA and LUCERNE: Two-Year Results from the Phase 3 Neovascular Age-Related Macular Degeneration Trials of Faricimab with Treat-and-Extend Dosing in Year 2.

PURPOSE: To evaluate 2-year efficacy, durability, and safety of the bispecific antibody faricimab, which inhibits both angiopoietin-2 and VEGF-A. DESIGN: TENAYA (ClinicalTrials.gov identifier, NCT03823287) and LUCERNE (ClinicalTrials.gov identifier, NCT03823300) were identically designed, randomized, double-masked, active comparator-controlled phase 3 noninferiority trials. PARTICIPANTS: Treatment-naive patients with neovascular age-related macular degeneration (nAMD) 50 years of age or older. METHODS: Patients were randomized (1:1) to intravitreal faricimab 6.0 mg up to every 16 weeks (Q16W) or aflibercept 2.0 mg every 8 weeks (Q8W). Faricimab fixed dosing based on protocol-defined disease activity at weeks 20 and 24 up to week 60, followed up to week 108 by a treat-and-extend personalized treatment interval regimen. MAIN OUTCOME MEASURES: Efficacy analyses included change in best-corrected visual acuity (BCVA) from baseline at 2 years (averaged over weeks 104, 108, and 112) and proportion of patients receiving Q16W, every 12 weeks (Q12W), and Q8W dosing at week 112 in the intention-to-treat population. Safety analyses included ocular adverse events (AEs) in the study eye through study end at week 112. RESULTS: Of 1326 patients treated across TENAYA/LUCERNE, 1113 (83.9%) completed treatment (n = 555 faricimab; n = 558 aflibercept). The BCVA change from baseline at 2 years was comparable between faricimab and aflibercept groups in TENAYA (adjusted mean change, +3.7 letters [95% confidence interval (CI), +2.1 to +5.4] and +3.3 letters [95% CI, +1.7 to +4.9], respectively; mean difference, +0.4 letters [95% CI, -1.9 to +2.8]) and LUCERNE (adjusted mean change, +5.0 letters [95% CI, +3.4 to +6.6] and +5.2 letters [95% CI, +3.6 to +6.8], respectively; mean difference, -0.2 letters [95% CI, -2.4 to +2.1]). At week 112 in TENAYA and LUCERNE, 59.0% and 66.9%, respectively, achieved Q16W faricimab dosing, increasing from year 1, and 74.1% and 81.2%, achieved Q12W or longer dosing. Ocular AEs in the study eye were comparable between faricimab and aflibercept groups in TENAYA (55.0% and 56.5% of patients, respectively) and LUCERNE (52.9% and 47.5% of patients, respectively) through week 112. CONCLUSIONS: Treat-and-extend faricimab treatment based on nAMD disease activity maintained vision gains through year 2, with most patients achieving extended dosing intervals. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Decreased FGF23 inhibits placental angiogenesis via the ERK1/2-EGR-1 signaling pathway in preeclampsia.

PURPOSE: This study aimed to investigate the expression of fibroblast growth factor 23 (FGF23) in pregnant women with preeclampsia and elucidate its role in promoting placental angiogenesis through the ERK1/2-EGR-1 signaling pathway. METHODS: Serum FGF23 levels were measured by ELISA in healthy pregnant women and patients with preeclampsia during the first, second, and third trimesters of pregnancy. Wound healing, Transwell, and tube formation assays were performed to investigate the effects of FGF23 on cell migration, invasion and tube formation. The expression of vascular endothelial growth factor A (VEGF-A) and its upstream signaling molecules, p-ERK, and EGR-1, in placental tissues was detected by RT-qPCR and western blotting. Additionally, the effect of FGF23 on VEGF-A, p-ERK, and EGR-1 expression was further explored in vitro. RESULTS: Serum FGF23 levels increased with gestational age. During the third trimester, the control group exhibited a more pronounced increase in FGF23 levels than the preeclampsia group. Administering exogenous FGF23 promoted trophoblast cell migration, invasion and enhanced tube formation in vascular endothelial cells. The expression levels of VEGF-A, p-ERK, and EGR-1 in the placental tissues were significantly lower in the preeclampsia group than in the control group. In vitro experiments confirmed that FGF23 up-regulated VEGF-A expression through the p-ERK/EGR-1 signaling pathway. CONCLUSION: The serum level of FGF23 decreased in pregnant women with preeclampsia, inhibiting the ERK1/2-EGR-1 pathway and resulting in decreased expression of VEGF-A, thereby inhibiting placental angiogenesis. This could be a potential mechanism involved in the progression of preeclampsia.

Impact of umbelliprenin-containing niosome nanoparticles on VEGF-A and CTGF genes expression in retinal pigment epithelium cells.

AIM: To investigate the impact of niosome nanoparticles carrying umbelliprenin (UMB), an anti-angiogenic and anti-inflammatory plant compound, on the expression of vascular endothelial growth factor (VEGF-A) and connective tissue growth factor (CTGF) genes in a human retinal pigment epithelium (RPE)-like retina-derived cell line. METHODS: UMB-containing niosomes were created, optimized, and characterized. RPE-like cells were treated with free UMB and UMB-containing niosomes. The IC50 values of the treatments were determined using an MTT assay. Gene expression of VEGF-A and CTGF was evaluated using real-time polymerase chain reaction after RNA extraction and cDNA synthesis. Niosomes' characteristics, including drug entrapment efficiency, size, dispersion index, and zeta potential were assessed. Free UMB had an IC50 of 96.2 µg/mL, while UMB-containing niosomes had an IC50 of 25 µg/mL. RESULTS: Treatment with UMB-containing niosomes and free UMB resulted in a significant reduction in VEGF-A expression compared to control cells (P=0.001). Additionally, UMB-containing niosomes demonstrated a significant reduction in CTGF expression compared to control cells (P=0.05). However, there was no significant reduction in the expression of both genes in cells treated with free UMB. CONCLUSION: Both free UMB and niosome-encapsulated UMB inhibits VEGF-A and CTGF genes expression. However, the latter demonstrates significantly greater efficacy, potentially due to the lower UMB dosage and gradual delivery. These findings have implications for anti-angiogenesis therapeutic approaches targeting age-related macular degeneration.

Is there a relationship between the first-day results of anti-VEGF therapy for macular edema due to vascular diseases and longterm outcomes? / Há alguma relação entre os resultados do primeiro dia de terapia anti-VEGF para edema macular devido a doenças vasculares e o resultado a longo prazo?

ABSTRACT Purpose: To evaluate early changes after the first antivascular endothelial growth factor injection for macular edema secondary to diabetic retinopathy and retinal vein occlusion and the relationship between longterm outcomes. Methods: The study enrolled patients who received anti-vascular endothelial growth factor injections for treatment-naive macular edema due to retinal vein occlusion and diabetic retinopathy. The central macular thickness was measured at baseline, post-injection day 1, week 2, and month 1, and at the last visit using spectral-domain optical coherence tomography. A good response was defined as a central macular thickness reduction of ≥10% on post-injection day 1. Patients were reassessed at the last visit with regard to treatment response on post-injection day 1 based on the favorable anatomic outcome defined as a central macular thickness <350 µm. Results: In total, 26 (44.8%) patients had macular edema-retinal vein occlusion and 32 (55.2%) had macular edema-diabetic retinopathy. The mean follow-up time was 24.0 (SD 8.5) months. A statistically significant decrease in the central macular thickness was observed in both patients with macular edema-retinal vein occlusion and macular edema-diabetic retinopathy after antivascular endothelial growth factor injection therapy (p<0.001 for both). All patients with macular edema-retinal vein occlusion were good responders at post-injection day 1. All nongood responders at post-injection day 1 belong to the macular edema-diabetic retinopathy group (n=16.50%). The rate of hyperreflective spots was higher in nongood responders than in good responders of the macular edema-diabetic retinopathy group (p=0.03). Of 42 (2.4%) total good responders, one had a central macular thickness >350 µm, whereas 5 (31.2%) of 16 total nongood responders had a central macular thickness >350 µm at the last visit (p=0.003). Conclusion: The longterm anatomical outcomes of macular edema secondary to retinal vein occlusion and diabetic retinopathy may be predicted by treatment response 1 day after antivascular endothelial growth factor injection.

RESUMO Objetivo: Avaliar as alterações precoces após a primeira injeção de anticorpos antifator de crescimento endotelial vascular (anti-VEGF) em casos de edema macular secundário à retinopatia diabética e oclusão da veia da retina e a relação entre essas alterações e o resultado a longo prazo. Métodos: Foram incluídos no estudo pacientes que receberam uma injeção de antifator de crescimento endotelial vascular para edema macular, virgem de tratamento e devido à oclusão da veia retiniana ou a retinopatia diabética. A espessura macular central foi medida no início do tratamento e no 1º dia, 2ª semana e 1º mês após a injeção, bem como na última visita, através de tomografia de coerência óptica de domínio espectral. Definiu-se uma "boa resposta" como uma redução ≥10% na espessura macular central no 1º dia após a injeção. Os pacientes foram reavaliados na última visita com relação à resposta ao tratamento no 1º dia após a injeção, com base em um resultado anatômico favorável, definido como uma espessura macular central <350 µm. Resultado: Foram registrados 26 (44,8%) pacientes com edema macular e oclusão da veia da retina e 32 (55,2%) com edema macular e retinopatia diabética. O tempo médio de acompanhamento foi de 24,0 meses (desvio-padrão de 8,5 meses). Foi observada uma diminuição estatisticamente significativa da espessura macular central após o tratamento antifator de crescimento endotelial vascular tanto em pacientes com edema macular e oclusão da veia retiniana quanto naqueles com edema macular e retinopatia diabética (p<0,001 para ambos). Todos os pacientes com edema macular e oclusão da veia retiniana responderam bem no 1º dia pós-injeção. Todos os que responderam mal no 1º dia pós-injeção pertenciam ao grupo com edema macular e retinopatia diabética (n=16,50%). A presença de manchas hiperrefletivas foi maior nos pacientes que responderam mal do que naqueles que tiveram boa resposta no grupo com edema macular e retinopatia diabética (p=0,03). Um dos 42 (2,4%) pacientes com boa resposta total teve espessura macular central >350 um, enquanto 5 (31,2%) do total de 16 pacientes com resposta ruim apresentaram espessura macular central >350 µm na última visita (p=0,003). Conclusão: O resultado anatômico de longo prazo do edema macular secundário à oclusão da veia retiniana e à retinopatia diabética pode ser previsto pela resposta ao tratamento no 1º dia após a injeção de antifator de crescimento endotelial vascular.

Expressions of vascular endothelial growth factor A, mucin-1, colony-stimulating factor-1, heparin-binding epidermal growth factor-like growth factor, and fibroblast growth factor 2 genes in the female reproductive tracts of women with ectopic pregnancy: A case-control study.

Background: Ectopic pregnancy (EP) is defined as embryo implantation in a location other than the uterine cavity. Objective: We aimed to evaluate the expression of several genes, which may play a role in EP, in the ampulla region of fallopian tubes and endometrial tissue of women with EP. Materials and Methods: In this case-control study, 5 women who underwent salpingectomy due to EP, comprised the 5 pseudo-pregnant women as a control group. These participants referred to the Royan Institute, Shariati, and Arash hospital, Tehran, Iran during 2019-2021. We evaluated the expressions of vascular endothelial growth factor A, mucin-1, colony-stimulating factor-1, heparin-binding epidermal growth factor-like growth factor (HBEGF), and fibroblast growth factor 2 genes in the fallopian tube and endometrium of EP cases by real-time polymerase chain reaction using specific primers. Results: The vascular endothelial growth factor expression was significantly higher in the ampulla region of the controls. However, no significant differences were observed in endometrial tissue. Assessments of colony-stimulating factor-1 and fibroblast growth factor 2 showed no significant differences between the case and control groups. HBEGF showed significantly higher expression in the ampulla region of EP cases, but no significant difference was observed in HBEGF expression in the endometrial tissues of the study groups. Mucin-1expression was significantly higher in both study regions of the EP cases. Conclusion: Our results have strongly suggested that these genes play important roles in proper implantation, and disruptions in their expression patterns could lead to EP. However, more studies are needed to confirm the current findings.

Faricimab Treat-and-Extend for Diabetic Macular Edema: Two-Year Results from the Randomized Phase 3 YOSEMITE and RHINE Trials.

PURPOSE: To evaluate the 2-year efficacy, durability, and safety of dual angiopoietin-2 and vascular endothelial growth factor (VEGF) A pathway inhibition with intravitreal faricimab according to a personalized treat-and-extend (T&E)-based regimen with up to every-16-week dosing in the YOSEMITE and RHINE (ClinicalTrials.gov identifiers, NCT03622580 and NCT03622593, respectively) phase 3 trials of diabetic macular edema (DME). DESIGN: Randomized, double-masked, noninferiority phase 3 trials. PARTICIPANTS: Adults with visual acuity loss (best-corrected visual acuity [BCVA] of 25-73 letters) due to center-involving DME. METHODS: Patients were randomized 1:1:1 to faricimab 6.0 mg every 8 weeks, faricimab 6.0 mg T&E (previously referred to as personalized treatment interval), or aflibercept 2.0 mg every 8 weeks. The T&E up to every-16-week dosing regimen was based on central subfield thickness (CST) and BCVA change. MAIN OUTCOME MEASURES: Included changes from baseline in BCVA and CST, number of injections, durability, absence of fluid, and safety through week 100. RESULTS: In YOSEMITE and RHINE (n = 940 and 951, respectively), noninferior year 1 visual acuity gains were maintained through year 2; mean BCVA change from baseline at 2 years (weeks 92, 96, and 100 average) with faricimab every 8 weeks (YOSEMITE and RHINE, +10.7 letters and +10.9 letters, respectively) or T&E (+10.7 letters and +10.1 letters, respectively) were comparable with aflibercept every 8 weeks (+11.4 letters and +9.4 letters, respectively). The median number of study drug injections was lower with faricimab T&E (YOSEMITE and RHINE, 10 and 11 injections, respectively) versus faricimab every 8 weeks (15 injections) and aflibercept every 8 weeks (14 injections) across both trials during the entire study. In the faricimab T&E arms, durability was improved further during year 2, with > 60% of patients receiving every-16-week dosing and approximately 80% receiving every-12-week or longer dosing at week 96. Almost 80% of patients who achieved every-16-week dosing at week 52 maintained every-16-week dosing without an interval reduction through week 96. Mean CST reductions were greater (YOSEMITE/RHINE weeks 92/96/100 average: faricimab every 8 weeks -216.0/-202.6 µm, faricimab T&E -204.5/-197.1 µm, aflibercept every 8 weeks -196.3/-185.6 µm), and more patients achieved absence of DME (CST < 325 µm; YOSEMITE/RHINE weeks 92-100: faricimab every 8 weeks 87%-92%/88%-93%, faricimab T&E 78%-86%/85%-88%, aflibercept every 8 weeks 77%-81%/80%-84%) and absence of intraretinal fluid (YOSEMITE/RHINE weeks 92-100: faricimab every 8 weeks 59%-63%/56%-62%, faricimab T&E 43%-48%/45%-52%, aflibercept every 8 weeks 33%-38%/39%-45%) with faricimab every 8 weeks or T&E versus aflibercept every 8 weeks through year 2. Overall, faricimab was well tolerated, with a safety profile comparable with that of aflibercept. CONCLUSIONS: Clinically meaningful visual acuity gains from baseline, anatomic improvements, and extended durability with intravitreal faricimab up to every 16 weeks were maintained through year 2. Faricimab given as a personalized T&E-based dosing regimen supports the role of dual angiopoietin-2 and VEGF-A inhibition to promote vascular stability and to provide durable efficacy for patients with DME. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Neutrophils are involved in early bone formation during midpalatal expansion.

OBJECTIVE: Midpalatal expansion (MPE) is routinely employed to treat transverse maxillary arch deficiency. Neutrophils are indispensable for recruiting bone marrow stromal cells (BMSCs) at the initial stage of bone regeneration. This study aimed to explore whether neutrophils participate in MPE and how they function during bone formation under mechanical stretching. MATERIALS AND METHODS: The presence and phenotype of neutrophils in the midpalatal suture during expansion were detected by flow cytometry and immunofluorescence staining. The possible mechanism of neutrophil recruitment and polarization was explored in vitro by exposing vascular endothelial cells (VECs) to cyclic tensile strain. RESULTS: The number of neutrophils in the distracted suture peaked on Day 3, and N2-type neutrophils significantly increased on Day 5 after force application. The depletion of circulatory neutrophils reduced bone volume by 43.6% after 7-day expansion. The stretched VECs recruited neutrophils via a CXCR2 mechanism in vitro, which then promoted BMSC osteogenic differentiation through the VEGFA/VEGFR2 axis. Consistently, these neutrophils showed higher expression of canonical N2 phenotype genes, including CD206 and Arg1. CONCLUSIONS: These results suggested that neutrophils participated in early bone formation during MPE. Based on these findings, we propose that stretched VECs recruited and polarized neutrophils, which, in turn, induced BMSC osteogenic differentiation.

A Retrospective Exploratory Analysis for Serum Extracellular Vesicles Reveals APRIL (TNFSF13), CXCL13, and VEGF-A as Prognostic Biomarkers for Neoadjuvant Chemotherapy in Triple-Negative Breast Cancer.

Neoadjuvant chemotherapy (NAC) is widely used as a standard treatment for early-stage triple-negative breast cancer (TNBC). While patients who achieve pathologic complete response (pCR) have a highly favorable outcome, patients who do not achieve pCR have variable prognoses. It is important to identify patients who are most likely to have poor survival outcomes to identify candidates for more aggressive therapeutic approaches after NAC. Many studies have demonstrated that cytokines and growth factors packaged into extracellular vesicles (EVs) have an essential role in tumor progression and drug resistance. In this study, we examined the role of serum-derived EV-associated cytokines as prognostic biomarkers for long-term outcomes in patients who underwent anthracycline-taxane-based NAC. We isolated extracellular vesicles from the serum of 190 TNBC patients who underwent NAC between 2015 and 2018 at Samsung Medical Center. EV-associated cytokine concentrations were measured with ProcartaPlex Immune Monitoring 65-plex panels. The prognostic value of EV-associated cytokines was studied. We found that patients with high EV_APRIL, EV_CXCL13, and EV_VEGF-A levels had shorter overall survival (OS). We further evaluated the role of these selected biomarkers as prognostic factors in patients with residual disease (RD) after NAC. Even in patients with RD, high levels of EV_APRIL, EV_CXCL13, and EV_VEGF-A were correlated with poor OS. In all subgroup analyses, EV_CXCL13 overexpression was significantly associated with poor overall survival. Moreover, multivariate analysis indicated that a high level of EV_CXCL13 was an independent predictor of poor OS. Correlation analysis between biomarker levels in EVs and serum showed that EV_VEGF-A positively correlated with soluble VEGF-A but not CXCL13. An elevated level of soluble VEGF-A was also associated with poor OS. These findings suggest that EV_APRIL, EV_CXCL13, and EV_VEGF-A may be useful in identifying TNBC patients at risk of poor survival outcomes after NAC.

[Expression of interleukin-37, vascular endothelial growth factor A, and transforming growth factor-ß1 and their correlation with T cells in children with primary immune thrombocytopenia]. / å„¿ç«¥åŽŸå‘å ç–«æ€§è¡€å°æ¿å‡å°‘ç—‡IL-37、VEGFA、TGF-ß1çš„è¡¨è¾¾åŠå ¶ä¸ŽTç»†èƒžçš„ç›¸å ³æ€§ç ”ç©¶.

OBJECTIVES: To investigate the expression of interleukin-37 (IL-37), vascular endothelial growth factor A (VEGFA), and transforming growth factor-ß1 (TGF-ß1) in children with primary immune thrombocytopenia (ITP) and their correlation with T cells. METHODS: A retrospective analysis was conducted on 45 children with ITP (ITP group) who were admitted to Handan Central Hospital from January 2020 to April 2022, and 30 healthy children who underwent physical examination during the same period were included as the healthy control group. The mRNA expression levels of IL-37, VEGFA, and TGF-ß1 and the levels of regulatory T cells (Treg) and helper T cells 17 (Th17) were measured before and after treatment, and the correlation between the mRNA expression levels of IL-37, VEGFA, and TGF-ß1 and the levels of Treg, Th17, and Treg/Th17 ratio were analyzed. RESULTS: Compared with the healthy control group, the ITP group had a significantly higher mRNA expression level of IL-37 and a significantly higher level of Th17 before and after treatment, as well as significantly lower mRNA expression levels of VEGFA and TGF-ß1 and significantly lower levels of Treg and Treg/Th17 ratio (P<0.05). After treatment, the ITP group had significant reductions in the mRNA expression level of IL-37 and the level of Th17 and significant increases in the mRNA expression levels of VEGFA and TGF-ß1 and the levels of Treg and Treg/Th17 ratio (P<0.05). Correlation analysis showed that in the ITP group, the mRNA expression levels of IL-37 and TGF-ß1 were negatively correlated with the levels of Treg and Treg/Th17 ratio (P<0.05) and were positively correlated with the level of Th17 (P<0.05) before and after treatment; the mRNA expression level of VEGFA was positively correlated with the levels of Treg and Treg/Th17 ratio (P<0.05) and was negatively correlated with the Th17 level (P<0.05) before and after treatment. CONCLUSIONS: Abnormal expression levels of IL-37, VEGFA, and TGF-ß1 may be observed in children with ITP, which is significantly associated with the imbalance of Treg/Th17 ratio. It is speculated that the cytokines such as IL-37, VEGFA, and TGF-ß1 may be involved in the development and progression of ITP or may become important potential targets for the treatment of children with ITP. Citation:Chinese Journal of Contemporary Pediatrics, 2023, 25(11): 1131-1136.

Inhibition of infantile hemangioma growth and promotion of apoptosis via VEGF/PI3K/Akt axis by 755-nm long-pulse alexandrite laser.

BACKGROUND: Infantile hemangioma (IH) is a common vascular tumor in female infants, which can lead to aesthetic issues and facial scarring. This study aimed to investigate the inhibitory effects and underlying mechanisms of 755 nm long-pulsed alexandrite laser on IH. METHODS: Hemangioma endothelial cells (HemECs) were exposed to 755 nm long-pulsed alexandrite laser to evaluate its impact on cell proliferation and apoptosis. A patient-derived xenograft model was established to assess the inhibitory effects of laser treatment on IH in vivo. RESULTS: In vitro, 755 nm long-pulsed alexandrite laser effectively suppressed the proliferation of HemECs and induced cell apoptosis. Laser treatment significantly inhibited the volume and weight of tumors, accompanied by significant downregulation of vascular endothelial growth factor A (VEGFA), vascular endothelial growth factor receptor 2 (VEGFR2), phosphatidylinositol 3-kinase (PI3K), and protein kinase B (Akt) expression levels in both hemangioma cells and tumors. Additionally, laser treatment resulted in the conversion of VEGFA165a to VEGFA165b. TUNEL staining demonstrated increased apoptosis in tumor cells after laser treatment, along with upregulation of cleaved caspase-3 and Bax, and downregulation of Bcl-2. CONCLUSION: In addition to the principle of selective photothermal decomposition, modulation of the VEGF/PI3K/Akt axis may serve as a potential mechanism for IH treatment using a long pulse-width 755 nm laser. This sheds valuable light on the molecular mechanisms underlying IH pathogenesis and potential therapeutic targets while providing a theoretical basis for the safe and efficient management of proliferative IH using laser therapy.

Neuron-specific deletion of VEGF or its receptor Flk-1 occludes the antidepressant-like effects of desipramine and fluoxetine in mice.

Vascular endothelial growth factor (VEGF) signaling is known to be involved in the antidepressant-like effects of conventional antidepressants, such as desipramine (DMI), a tricyclic antidepressant, and fluoxetine (FLX), a selective serotonin reuptake inhibitor; however, the precise role of neuronal VEGF signaling in mediating these effects remains unclear. Using mice with excitatory neuron-specific deletion of VEGF and its receptor, fetal liver kinase 1 (Flk-1) in the forebrain, we examined the effects of forebrain excitatory neuron-specific deletion of VEGF or Flk-1 on the antidepressant-like effects of repeated DMI and chronic FLX administration in the forced swim test (FST). Repeated intraperitoneal (i.p.) injections of DMI (10, 10, and 20 mg/kg at 24, 4, and 1 h before the FST, respectively) significantly decreased immobility in control mice; however, this effect was completely blocked in mice with neuron-specific VEGF or Flk-1 deletion. Although chronic treatment with FLX (18 mg/kg/day, i.p.) did not impact immobility in control mice 1 day after the 22nd injection, immobility was significantly reduced 1 day after the preswim and the 23rd FLX injection. However, in mice with neuron-specific Flk-1 deletion, chronic FLX treatment significantly increased immobility in the preswim and failed to produce antidepressant-like effects. Collectively, these findings indicate that neuronal VEGF-Flk-1 signaling contributes to the antidepressant-like actions of conventional antidepressants.

Vasoregulatory Autoantibodies and Clinical Outcome After Ischemic Stroke-PROSCIS-B.

BACKGROUND: Vasoregulatory autoantibodies including autoantibodies targeting G-protein-coupled receptors might play a functional role in vascular diseases. We investigated the impact of vasoregulatory autoantibodies on clinical outcome after ischemic stroke. METHODS AND RESULTS: Data were used from the PROSCIS-B (Prospective Cohort With Incident Stroke-Berlin). Autoantibody-targeting receptors such as angiotensin II type 1 receptor (AT1R), endothelin-1 type A receptor, complement factor-3 and -5 receptors, vascular endothelial growth factor receptor-1 and -2, vascular endothelial growth factor A and factor B were measured. We explored associations of high antibody levels with (1) poor functional outcome defined as modified Rankin Scale >2 or Barthel Index <60 at 1 year after stroke, (2) Barthel Index scores over time using general estimating equations, and (3) secondary vascular events (recurrent stroke, myocardial infarction) or death up to 3 years using Cox proportional hazard models. We included 491 patients with ischemic stroke with data on autoantibody levels and outcome. In models adjusted for demographics and vascular risk factors, high autoantibody concentrations (quartile 4) targeting complement factor C3a receptor, vascular endothelial growth factor receptor-2, and vascular endothelial growth factor B were associated with poor functional outcome at 1 year: (odds ratio, 2.0 [95% CI, 1.1-3.6]; odds ratio, 1.8 [95% CI, 1.1-3.2]; and odds ratio, 2.1 [95% CI, 1.2-3.6], respectively) and with lower Barthel Index scores over 3 years (complement factor C3a receptor: adjusted ß=-3.3 [95% CI, -5.7 to -0.5]; VEGF-B: adjusted ß=-2.4 [95% CI, -4.8 to -0.06]). Patients with high autoantibody levels were not at higher risk for secondary vascular events or death. CONCLUSIONS: High levels of autoantibodies against vascular endothelial growth factor receptor-2, vascular endothelial growth factor B, and complement factor C3a receptor measured are associated with poor functional outcome after stroke but not with recurrent vascular events or death. REGISTRATION: URL: https://www.clinicaltrials.gov; Unique identifier: NCT01363856.

Estrogen receptor-related receptor γ uppresses hypoxia-induced angiogenesis by regulating VEGFA in endometrial cancer.

OBJECTIVE: Estrogen receptor-related receptor γ (ERRγ), is implicated in cancer cell proliferation and metastasis. The function of ERRγ in tumor angiogenesis, however, is to be revealed. This study was designed to elaborate the regulatory effect of ERRγ on angiogenesis in endometrial cancer (EC). METHODS: Immunohistochemistry (IHC) was adopted to determine the protein expression of ERRγ, VEGFA, CD31 and hypoxia-inducible factor-1 (HIF-1) in tumor tissues. HEC-1A cells stably expressing ERRγ were established bytransfection, and then an endothelial cell tube formation assay was performed. CCK-8 assay was employed for cell viability, and wound healing assay for cell migration ability. Besides, western blot, ELISA and qRT-PCR were used to examine the VEGFA expression. After hypoxia treatment of ERRγ overexpressing HEC-1A cells, the ERRγ expression and VEGFA expression were determined by western blot. Finally, EC xenografts in nude mice were constructed by subcutaneous injection of ERRγ stably expressing HEC-1A cells and control HEC-1A cells. RESULTS: IHC results revealed a negative correlation between the expression of ERRγ and VEGFA in EC tissues. ERRγ overexpression significantly decreased the level of HIF-1 in tumor tissue of nude mice. ERRγ overexpression down-regulated inhibited angiogenesis capability and inhibited the proliferation and migration of HEC-1A cells. Furthermore, ERRγ expression was suppressed under the condition of hypoxia while restoration of ERRγ partially inhibited hypoxia-induced VEGFA expression in HEC-1A cells. CONCLUSIONS: ERRγ is an angiogenesis suppressor and involved in hypoxia-induced VEGFA expression in EC. Hence, ERRγ might be a promising antiangiogenic target for human EC.