Dual crosslinking silk fibroin/pectin-based bioink development and the application on neural stem/progenitor cells spheroid laden 3D bioprinting.

The human nervous system is an incredibly intricate physiological network, and neural cells lack the ability to repair and regenerate after a brain injury. 3-dimensional (3D) bioprinting technology offers a promising strategy for constructing biomimetic organ constructs and in vitro brain/disease models. The bioink serves as a pivotal component that emulates the microenvironment of biomimetic construct and exerts a profound influence on cellular behaviors. In this study, a series of mechanically adjustable and dual crosslinking bioinks were developed using photocrosslinkable methacrylated silk fibroin (SilMA) in combination with the ionic crosslinking material, pectin, or pectin methacryloyl (PecMA) with silk fibroin (SF) supplementation. SilMA/pectin exhibited superior properties, with SilMA providing biocompatibility and adjustable mechanical properties, while the addition of pectin enhanced printability. The porous structure supported neural cell growth, and 15 % SilMA/0.5 % pectin bioinks displayed excellent printability and shape fidelity. Neural stem/progenitor cells (NSPCs)-loaded bioinks were used to construct a 3D brain model, demonstrating sustained vitality and high neuronal differentiation without the need for growth factors. The SilMA/pectin bioinks demonstrated adjustable mechanical properties, favorable biocompatibility, and an environment highly conducive to neural induction, offering an alternative approach for neural tissue engineering applications or in vitro brain models.

Genetic Deletion of ß-Arrestin 2 From the Subfornical Organ and Other Periventricular Nuclei in the Brain Alters Fluid Homeostasis and Blood Pressure.

BACKGROUND: ANG (angiotensin II) elicits dipsogenic and pressor responses via activation of the canonical Gαq (G-protein component of the AT1R [angiotensin type 1 receptor])-mediated AT1R in the subfornical organ. Recently, we demonstrated that ARRB2 (ß-arrestin 2) global knockout mice exhibit a higher preference for salt and exacerbated pressor response to deoxycorticosterone acetate salt. However, whether ARRB2 within selective neuroanatomical nuclei alters physiological responses to ANG is unknown. Therefore, we hypothesized that ARRB2, specifically in the subfornical organ, counterbalances maladaptive dipsogenic and pressor responses to the canonical AT1R signaling. METHODS: Male and female Arrb2FLOX mice received intracerebroventricular injection of either adeno-associated virus (AAV)-Cre-GFP (green fluorescent protein) to induce brain-specific deletion of ARRB2 (Arrb2ICV-Cre). Arrb2FLOX mice receiving ICV-AAV-GFP were used as control (Arrb2ICV-Control). Infection with ICV-AAV-Cre primarily targeted the subfornical organ with few off targets. Fluid intake was evaluated using the 2-bottle choice paradigm with 1 bottle containing water and 1 containing 0.15 mol/L NaCl. RESULTS: Arrb2ICV-Cre mice exhibited a greater pressor response to acute ICV-ANG infusion. At baseline conditions, Arrb2ICV-Cre mice exhibited a significant increase in saline intake compared with controls, resulting in a saline preference. Furthermore, when mice were subjected to water-deprived or sodium-depleted conditions, which would naturally increase endogenous ANG levels, Arrb2ICV-Cre mice exhibited elevated saline intake. CONCLUSIONS: Overall, these data indicate that ARRB2 in selective cardiovascular nuclei in the brain, including the subfornical organ, counterbalances canonical AT1R responses to both exogenous and endogenous ANG. Stimulation of the AT1R/ARRB axis in the brain may represent a novel strategy to treat hypertension.

Long-term outcomes of inferomedial orbital wall decompression in Graves' ophthalmopathy in an Asian population: A 30-year retrospective study.

PURPOSE: This study aims to evaluate the long-term outcomes of inferomedial orbital wall decompression (IMOD) in Graves' ophthalmopathy (GO). METHODS: A retrospective review of 422 eyes of 226 patients with GO-related cosmetically disfiguring proptosis (COS), dysthyroid optic neuropathy (DON), or exposure keratopathy (EXP) who received IMOD from 1989 to 2020 was conducted. Hertel value (HE) and corrected visual acuity (CVA) were evaluated at baseline and regularly thereafter. Proptosis recurrence, diplopia, and adjuvant surgeries were assessed. Complete success was defined as proptosis reduction over 2 mm without recurrence and improved diplopia. Partial success was defined as proptosis reduction without recurrence but with persistent or new-onset diplopia, and failure as proptosis recurrence. RESULTS: After follow-up for 40.1 ± 39.6 months (range, 6-239 months), 236 eyes (55.9%) achieved complete success, 175 eyes (41.5%) achieved partial success, and 11 eyes (2.6%) had failure. Significant CVA improvement was observed in both DON and COS groups (P < 0.001 and P = 0.045, respectively). Average proptosis reduction was 5.2 ± 2.2 mm, with a significant increase of 0.5 mm after long-term follow-up (P < 0.001). Incidence of proptosis recurrence was 2.6%. Surgical success rate of COS group was positively correlated with preoperative HE. CONCLUSION: IMOD demonstrated excellent success rate in all three subgroups with a low proptosis recurrence rate and improved visual outcome in both DON and COS groups. Delayed proptosis reduction effect of IMOD may result in delayed-onset asymmetry, especially in the unilateral group.

Evaluation the Effect of Sonodynamic Therapy with 5-Aminolevulinic Acid and Sodium Fluorescein by Preclinical Animal Study.

Sonodynamic therapy (SDT) is a novel tumor treatment that combines biosafe sonosensitizers and noninvasive focused ultrasound to eradicate solid tumors. Sonosensitizers such as 5-aminolevulinic acid and fluorescein have great potential in tumor treatment. Here, rodent subcutaneous and brain tumor models were used to evaluate the treatment effect of both 5-ALA- and fluorescein-mediated SDT. The subcutaneous tumor growth rates of both SDT groups were significantly inhibited compared with that of the control groups. For intracranial tumors, 5-ALA-SDT treatment significantly inhibited brain tumor growth, while fluorescein-SDT exerted no therapeutic effect in animals. The distribution of fluorescein in the brain tumor region underwent further assessment. Seven days post tumor implantation, experimental animals received fluorescein and were sacrificed for brain specimen collection. Analysis of the dissected brains revealed no fluorescence signals, indicating an absence of fluorescein accumulation in the early-stage glioma tissue. These data suggest that the fluorescein-SDT treatment response is closely related to the amount of accumulated fluorescein. This study reports the equivalent effects of 5-ALA and fluorescein on the treatment of somatic tumors. For orthotopic brain tumor models, tumor vascular permeability should be considered when choosing fluorescein as a sonosensitizer. In conclusion, both fluorescein and 5-ALA are safe and effective SDT sonosensitizers, and the tumor microenvironment and pathologic type should be considered in the selection of adequate sonosensitizers.

Minimally invasive endoscopic evacuation with the novel, portable Axonpen neuroendoscopic system for spontaneous intracerebral hemorrhage.

Minimally invasive surgeries have shown potential to improve mortality and clinical outcomes of spontaneous intracerebral hemorrhage (ICH). The present study assessed the first-in-human outcomes of a novel, portable neuroendoscopic system for ICH evacuation at our single center. This neuroendoscopic system integrates real-time visualization into a handpiece which has controllable suction, irrigation, and coagulation to allow a neurosurgeon to conduct minimally invasive ICH evacuation independently with bimanual dexterity. Pre- and postoperative data of ten patients who had spontaneous basal ganglia hemorrhage (mean: 46.5 ± 12.2 mL) and underwent evacuation with the specified neuroendoscopic system were collected prospectively. The mean time to receive surgery was 12.1 ± 7.6 h. Mean operative time was 3.4 ± 0.9 h. The mean hematoma volume decreased to 6.0 ± 3.9 mL at postoperative 6 h, resulting in a mean volume reduction of 86.0 ± 11.2% (P = 0.005). The median length of intensive care unit stay was 3 days (IQR, 3-4 days). At discharge, the median Glasgow Coma Scale (GCS) score significantly improved to 11.5 (IQR, 11-15; P = 0.016), and the median modified Rankin Scale (mRS) score was 4 (IQR, 4-5). Six patients (60%) showed a favorable mRS score of ≤ 3 on their last return visit. Neither death nor rebleeding occurred during the follow-up periods. Integrated design of the innovative device is valuable to optimize minimally invasive endoscopic ICH evacuation procedure. Further studies are needed to clarify long-term benefits from such type of the innovative device to early intervention of ICH.

Krüppel-like factor 4 in transcriptional control of the three unique isoforms of Agouti-related peptide in mice.

Agouti-related peptide (AgRP/Agrp) within the hypothalamic arcuate nucleus (ARC) contributes to the control of energy balance, and dysregulated Agrp may contribute to metabolic adaptation during prolonged obesity. In mice, three isoforms of Agrp are encoded via distinct first exons. Agrp-A (ENSMUST00000005849.11) contributed 95% of total Agrp in mouse ARC, whereas Agrp-B (ENSMUST00000194654.2) dominated in placenta (73%). Conditional deletion of Klf4 from Agrp-expressing cells (Klf4Agrp-KO mice) reduced Agrp mRNA and increased energy expenditure but had no effects on food intake or the relative abundance of Agrp isoforms in the ARC. Chronic high-fat diet feeding masked these effects of Klf4 deletion, highlighting the context-dependent contribution of KLF4 to Agrp control. In the GT1-7 mouse hypothalamic cell culture model, which expresses all three isoforms of Agrp (including Agrp-C, ENSMUST00000194091.6), inhibition of extracellular signal-regulated kinase (ERK) simultaneously increased KLF4 binding to the Agrp promoter and stimulated Agrp expression. In addition, siRNA-mediated knockdown of Klf4 reduced expression of Agrp. We conclude that the expression of individual isoforms of Agrp in the mouse is dependent upon cell type and that KLF4 directly promotes the transcription of Agrp via a mechanism that is superseded during obesity.NEW & NOTEWORTHY In mice, three distinct isoforms of Agouti-related peptide are encoded via distinct first exons. In the arcuate nucleus of the hypothalamus, Krüppel-like factor 4 stimulates transcription of the dominant isoform in lean mice, but this mechanism is altered during diet-induced obesity.

Focused ultrasound combined with radiotherapy for malignant brain tumor: a preclinical and clinical study.

INTRODUCTION: Blood-brain barrier (BBB) remains to be the major obstacle to conquer in treating patients with malignant brain tumors. Radiation therapy (RT), despite being the mainstay adjuvant modality regardless of BBB, the effect of radiation induced cell death is hindered by the hypoxic microenvironment. Focused ultrasound (FUS) combined with systemic microbubbles has been shown not only to open BBB but also potentially increased regional perfusion. However, no clinical study has investigated the combination of RT with FUS-BBB opening (RT-FUS). METHODS: We aimed to provide preclinical evidence of RT-FUS combination in GBM animal model, and to report an interim analysis of an ongoing single arm, prospective, pilot study (NCT01628406) of combining RT-FUS for recurrent malignant high grade glioma patients, of whom re-RT was considered for disease control. In both preclinical and clinical studies, FUS-BBB opening was conducted within 2 h before RT. Treatment responses were evaluated by objective response rate (ORR) using magnetic resonance imaging, progression free survival, and overall survival, and adverse events (AE) in clinical study. Survival analysis was performed in preclinical study and descriptive analysis was performed in clinical study. RESULTS: In mouse GBM model, the survival analysis showed RT-FUS (2 Gy) group was significantly longer than RT (2 Gy) group and control, but not RT (5 Gy) group. In the pilot clinical trial, an interim analysis of six recurrent malignant high grade glioma patients underwent a total of 24 RT-FUS treatments was presented. Three patients had rapid disease progression at a mean of 33 days after RT-FUS, while another three patients had at least stable disease (mean 323 days) after RT-FUS with or without salvage chemotherapy or target therapy. One patient had partial response after RT-FUS, making the ORR of 16.7%. There was no FUS-related AEs, but one (16.7%) re-RT-related grade three radiation necrosis. CONCLUSION: Reirradiation is becoming an option after disease recurrence for both primary and secondary malignant brain tumors since systemic therapy significantly prolongs survival in cancer patients. The mechanism behind the synergistic effect of RT-FUS in preclinical model needs further study. The clinical evidence from the interim analysis of an ongoing clinical trial (NCT01628406) showed a combination of RT-FUS was safe (no FUS-related adverse effect). A comprehensive analysis of radiation dosimetry and FUS energy distribution is expected after completing the final recruitment.

Mitochondrial-targeted antioxidant attenuates preeclampsia-like phenotypes induced by syncytiotrophoblast-specific Gαq signaling.

Syncytiotrophoblast stress is theorized to drive development of preeclampsia, but its molecular causes and consequences remain largely undefined. Multiple hormones implicated in preeclampsia signal via the Gαq cascade, leading to the hypothesis that excess Gαq signaling within the syncytiotrophoblast may contribute. First, we present data supporting increased Gαq signaling and antioxidant responses within villous and syncytiotrophoblast samples of human preeclamptic placenta. Second, Gαq was activated in mouse placenta using Cre-lox and DREADD methodologies. Syncytiotrophoblast-restricted Gαq activation caused hypertension, kidney damage, proteinuria, elevated circulating proinflammatory factors, decreased placental vascularization, diminished spiral artery diameter, and augmented responses to mitochondrial-derived superoxide. Administration of the mitochondrial-targeted antioxidant Mitoquinone attenuated maternal proteinuria, lowered circulating inflammatory and anti-angiogenic mediators, and maintained placental vascularization. These data demonstrate a causal relationship between syncytiotrophoblast stress and the development of preeclampsia and identify elevated Gαq signaling and mitochondrial reactive oxygen species as a cause of this stress.

Predictors for delayed awakening in adult glioma patients receiving awake craniotomy under monitored anesthesia care.

PURPOSE: Delayed awakening after anesthetic discontinuation during awake craniotomy is associated with somnolence during functional brain mapping. However, predictors of delayed awakening in patients receiving monitored anesthesia care for awake craniotomy are unknown. METHODS: This retrospective cohort study analyzed 117 adult patients with supratentorial glioma in or near eloquent areas who received monitored anesthesia care for awake craniotomy between July 2020 and January 2023 at Linkou Chang Gung Memorial Hospital. These patients were divided into two groups according to their time to awakening (ability to speak their names) after propofol cessation: longer or shorter than 20 min (median duration). Because propofol was solely used anesthetic from skin incision to dural opening, parameters in Schnider model for propofol target-controlled infusion, such as age, sex, and BMI, were adjusted or propensity-matched to compare their anesthetic, surgical, and histopathological profiles. RESULTS: After propensity-matched comparisons of age and BMI, significant predictors of delayed awakening included IDH1 wild-type tumors and repeated craniotomies. Subgroup analysis revealed that older age and larger T2 volume were predictors in patients undergoing the first craniotomy, while lower preoperative Karnofsky performance scale scores and depression were predictors in repeated craniotomy cases. Delayed awakening was also associated with somnolence and a lower gross total resection rate. CONCLUSION: Our retrospective analysis of patients receiving monitored anesthesia care for awake craniotomy revealed that delayed awakening after propofol discontinuation occurred more often in patients with IDH1 wild-type tumors and repeated craniotomies. Also, delayed awakening was associated with somnolence during functional mapping and a lower gross total resection rate.

Primary Graviceptive System and Astasia: A Case Report and Literature Review.

Astasia refers to the inability to maintain upright posture during standing, despite having full motor strength. Impairment of the vestibulocerebellar pathway, graviceptive system, and cingulate motor area have been proposed to be related to astasia. However, the responsible neural pathways remain unclear. We hypothesize that there is a common neural network behind astasia. To test the hypothesis, we reviewed all reported cases with astasia, including ours, and focused on the correlation between anatomical destruction and symptom presentation. A total of 26, including ours, non-psychogenic astasia patients were identified in the English literature. Seventy-three percent of them were associated with other neurologic symptoms and sixty-two percent of reported lesions were on the right side. Contralateral lateropulsion was very common, followed by retropulsion, when describing astasia. Infarction (54%) was the most reported cause. The thalamus (65%) was the most reported location. Infarctions were the fastest to recover (mean: 10.6 days), while lesions at the brainstem needed a longer time (mean: 61.6 days). By combining the character of lateropulsion in astasia and the presentation of an interrupted graviceptive system, we concluded that the primary graviceptive system may be the common neural network behind astasia. Future studies on astasia should focus on the pathological changes in the perception of verticality in the visual world and the body.

Adaptive focal lengths in white light focusing Fresnel lenses enabled by reflective-type and phase-only spatial light modulator.

Fresnel zone plates (FZPs) are widely used in integrated optical systems to meet new cutting-edge demands for photonic integration and device miniaturizing. However, their use in applications of cross-scale fabrication still faces several obstacles, such as low efficiency, fixed focal length, single wavelength, large size, and complicated fabrication. Here, we first examine a novel adaptive focal length in white light focusing by using reflective-type and phase-only spatial light modulator (RLC-SLM) based on a liquid crystal on silicon. The device achieves a maximum diffraction efficiency of approximately 38% at primary focal points of binary phase-type FZPs throughout the visible range (red, green, and blue wavelengths). The RLC-SLM focuses the light of the desired wavelength while other sources are defocused. White light focusing and color separation are demonstrated by sequentially and additively switching different FZPs. These recent advances show that optically tunable FRZs are promising potential candidates to enhance adaptive camera systems, microscopes, holograms, and portable and wearable devices, thereby opening up novel possibilities in optical communications and sensing.

Neuropsychological impairment in primary malignant brain tumor patients with awake craniotomy: a hospital-based registration study.

PURPOSE: Neuroplasticity is an ability to maintain neural circuit function when facing damages. It is one of the reasons that making brain tumors notorious. Therefore, we evaluated the characteristics of patients with primary brain tumors, compared neuropsychological deficits between patients who had awake craniotomy with left- or right-sided tumors, and analyzed the association between white matter tracts and neuropsychological deficits in patients with right-sided tumors. METHODS: Using the registration dataset of Chang Gung Memory Hospital between 2014 and 2020, this study included a total of 698 adult patients who received craniotomy for primary brain tumors (538 of conventional craniotomy; 160 of awake craniotomy). Neuropsychological assessments were arranged in patients as preoperative evaluation for awake craniotomies. RESULTS: A lower proportion of right-sided tumors was noted in patient who had awake craniotomy than those who had conventional craniotomy (33.8% and 51.5%, p < 0.001). In awake craniotomy, 88.7% of patients with left-sided tumors and 77.8% of patients with right-sided tumors had neuropsychological impairment. Patients with left-sided tumors had worse preoperative performance compared to those with right-sided tumors in global function (36.2% and 8.0%, p < 0.001), language domain (57.6% and 22.2%, p < 0.001), and attention (36.0% and 18.5%, p = 0.02). Furthermore, in those with right-sided low-grade gliomas, patients involving pathway of superior longitudinal fasciculus (SLF) I had a higher risk of deficits than those without involvement in verbal memory (p = 0.001, Odd ratio = 11.2, 95% CI = 1.8 ~ 71.4) and visual memory (p = 0.048, Odd ratio = 10.5, 95% CI = 1.0 ~ 111). CONCLUSION: In awake craniotomy, patients with left-sided brain tumors had worse cognitive function than those with right-sided tumors in terms of global function, language, and attention. 77% of patients with right-sided tumors had neuropsychological impairment. Therefore, a comprehensive neuropsychological evaluation and awake craniotomy are necessary for patients with brain tumors.

Technological process optimization and measurement of image quality of the electrically bifocal metalens.

This Letter describes the design procedure and process optimization of the electrically bifocal metalens. In our design, horizontal and vertical polarization is manipulated by applying a suitable voltage to a twisted nematic liquid crystal (TN-LC) cell. Each nanostructure is designed to be a rectangular prism, making different polarizations of light experience various phase delays, thus causing bi-focus. We selected lithographical methods to fabricate our metalens because of the minimum physical size, which can be as small as 50 nm, and the maximum aspect ratio, which is as high as 15. Furthermore, to increase the tolerance and make the sidewall vertical and smooth, we coated different characteristics of photoresist sensitivity to the upper and lower layers. After the development, the mushroom-type photoresist makes Ni easier to strip while in the lift-off process, thus increasing the quality of the whole metalens. Our experiment shows that the focal lengths and focusing efficiencies corresponding to the two polarizations are similar to the simulation results. The proposed electrically modulated bifocal metalens can be utilized in different applications and combined with other optical components.

CAN008 prolongs overall survival in patients with newly diagnosed GBM characterized by high tumor mutational burden.

BACKGROUND: A previous phase 1 dose-escalation study in Taiwan indicated CAN008 (asunercept) with standard concurrent chemoradiotherapy (CCRT) improved progression-free survival (PFS) in newly diagnosed glioblastoma (GBM) patients. This study evaluates the efficacy of CAN008 in promoting overall survival (OS) and identifies genetic alterations associated with treatment responses. METHODS: We compared OS of 5-year follow-ups from 9 evaluable CAN008 cohort patients (6 received high-dose and 3 received low-dose) to a historical Taiwanese GBM cohort with 164 newly diagnosed patients. CAN008 treatment response-associated genetic alterations were identified by whole-exome sequencing and comparing variant differences between response groups. Associations among patient survival, tumor mutational burden (TMB), and genetic alterations were analyzed using CAN008 cohort and TCGA-GBM dataset. RESULTS: OS for high-dose CAN008 patients at 2 and 5 years was 83% and 67%, respectively, and 40.1% and 8.8% for the historical GBM cohort, respectively. Better OS was observed in the high-dose CAN008 cohort (without reaching the median survival) than the historical GBM cohort (median OS: 20 months; p=0.0103). Five high-dose CAN008 patients were divided into good and poor response groups based on their PFS. A higher variant count and TMB were observed in good response patients, whereas no significant association was observed between TMB and patient survival in the newly diagnosed TCGA-GBM dataset, suggesting TMB may modulate patient CAN008 response. CONCLUSION: CAN008 combined with standard CCRT treatment prolonged the PFS and OS of newly diagnosed GBM patients compared to standard therapy alone. Higher treatment efficacy was associated with higher TMB.

Clinical outcome analysis of non-small cell lung cancer patients with brain metastasis receiving metastatic brain tumor resection surgery: a multicenter observational study.

Brain metastasis is most common in primary non-small cell lung cancer (NSCLC), and some patients require neurosurgical resection for intracranial disease control. Because advances in systemic therapies for metastatic NSCLC have been developed in the past decade, we aimed to analyze and determine clinical factors associated with the postresection survival of NSCLC patients with brain metastasis who underwent neurosurgery followed by systemic therapy. Between January 2017 and December 2021, data for 93 NSCLC patients with brain metastasis treated with neurosurgery followed by systemic therapy at Linkou, Kaohsiung and Chiayi Chang Gung Memorial Hospitals were retrospectively retrieved for analysis. For all study patients, median postresection survival was 34.36 months (95% confidence interval (CI), 28.97-39.76), median brain metastasis (BM)-free survival was 26.90 months (95% CI, 22.71-31.09), and overall survival (OS) was 41.13 months (95% CI, 34.47-47.52). In multivariate analysis, poor performance status (Eastern Cooperative Oncology Group performance status (ECOG PS) ≥2) and concurrent liver metastasis were identified as independent unfavorable factors associated with significantly shortened postresection survival (P<0.001). The histological type adenocarcinoma was associated with significantly longer postresection survival (P = 0.001). The median postresection survival for adenocarcinoma and nonadenocarcinoma patients was 36.23 and 10.30 months, respectively (hazard ratio (HR) = 0.122; 95% CI, 0.035-0.418; P<0.001); that for patients with and without concurrent liver metastasis was 11.43 and 36.23 months, respectively (HR = 22.18; 95% CI, 5.827-84.459; P<0.001). Patients with preserved ECOG PS, adenocarcinoma histology type and no concurrent liver metastasis appeared to have better postresection survival than nonadenocarcinoma patients. Our results provide counseling and decision-making references for neurosurgery feasibility in NSCLC patients with brain metastasis.

Genetic Ablation of Prorenin Receptor in the Rostral Ventrolateral Medulla Influences Blood Pressure and Hydromineral Balance in Deoxycorticosterone-Salt Hypertension.

Non-enzymatic activation of renin via its interaction with prorenin receptor (PRR) has been proposed as a key mechanism of local renin-angiotensin system (RAS) activation. The presence of renin and angiotensinogen has been reported in the rostral ventrolateral medulla (RVLM). Overactivation of bulbospinal neurons in the RVLM is linked to hypertension (HTN). Previous studies have shown that the brain RAS plays a role in the pathogenesis of the deoxycorticosterone (DOCA)-salt HTN model. Thus, we hypothesized that PRR in the RVLM is involved in the local activation of the RAS, facilitating the development of DOCA-salt HTN. Selective PRR ablation targeting the RVLM (PRRRVLM-Null mice) resulted in an unexpected sex-dependent and biphasic phenotype in DOCA-salt HTN. That is, PRRRVLM-Null females (but not males) exhibited a significant delay in achieving maximal pressor responses during the initial stage of DOCA-salt HTN. Female PRRRVLM-Null subsequently showed exacerbated DOCA-salt-induced pressor responses during the "maintenance" phase with a maximal peak at 13 d on DOCA-salt. This exacerbated response was associated with an increased sympathetic drive to the resistance arterioles and the kidney, exacerbated fluid and sodium intake and output in response to DOCA-salt, and induced mobilization of fluids from the intracellular to extracellular space concomitant with elevated vasopressin. Ablation of PRR suppressed genes involved in RAS activation and catecholamine synthesis in the RVLM but also induced expression of genes involved in inflammatory responses. This study illustrates complex and sex-dependent roles of PRR in the neural control of BP and hydromineral balance through autonomic and neuroendocrine systems. Graphical abstract.

Structure and Function of RhoBTB1 Required for Substrate Specificity and Cullin-3 Ubiquitination.

We identified Rho-related BTB domain containing 1 (RhoBTB1) as a key regulator of phosphodiesterase 5 (PDE5) activity, and through PDE5, a regulator of vascular tone. We identified the binding interface for PDE5 on RhoBTB1 by truncating full-length RhoBTB1 into its component domains. Co-immunoprecipitation analyses revealed that the C-terminal half of RhoBTB1 containing its two BTB domains and the C-terminal domain (B1B2C) is the minimal region required for PDE5 recruitment and subsequent proteasomal degradation via Cullin-3 (CUL3). The C-terminal domain was essential in recruiting PDE5 as constructs lacking this region could not participate in PDE5 binding or proteasomal degradation. We also identified Pro353 and Ser363 as key amino acid residues in the B1B2C region involved in CUL3 binding to RhoBTB1. Mutation of either of these residues exhibited impaired CUL3 binding and PDE5 degradation, although the binding to PDE5 was preserved. Finally, we employed ascorbate peroxidase 2 (APEX2) proximity labeling using a B1B2C-APEX2 fusion protein as bait to capture unknown RhoBTB1 binding partners. Among several B1B2C-binding proteins identified and validated, we focused on SET domain containing 2 (SETD2). SETD2 and RhoBTB1 directly interacted, and the level of SETD2 increased in response to pharmacological inhibition of the proteasome or Cullin complex, CUL3 deletion, and RhoBTB1-inhibition with siRNA. This suggests that SETD2 is regulated by the RhoBTB1-CUL3 axis. Future studies will determine whether SETD2 plays a role in cardiovascular function.

Acrolein produced by glioma cells under hypoxia inhibits neutrophil AKT activity and suppresses anti-tumoral activities.

Acrolein, which is the most reactive aldehyde, is a byproduct of lipid peroxidation in a hypoxic environment. Acrolein has been shown to form acrolein-cysteine bonds, resulting in functional changes in proteins and immune effector cell suppression. Neutrophils are the most abundant immune effector cells in circulation in humans. In the tumor microenvironment, proinflammatory tumor-associated neutrophils (TANs), which are termed N1 neutrophils, exert antitumor effects via the secretion of cytokines, while anti-inflammatory neutrophils (N2 neutrophils) support tumor growth. Glioma is characterized by significant tissue hypoxia, immune cell infiltration, and a highly immunosuppressive microenvironment. In glioma, neutrophils exert antitumor effects early in tumor development but gradually shift to a tumor-supporting role as the tumor develops. However, the mechanism of this anti-to protumoral switch in TANs remains unclear. In this study, we found that the production of acrolein in glioma cells under hypoxic conditions inhibited neutrophil activation and induced an anti-inflammatory phenotype by directly reacting with Cys310 of AKT and inhibiting AKT activity. A higher percentage of cells expressing acrolein adducts in tumor tissue are associated with poorer prognosis in glioblastoma patients. Furthermore, high-grade glioma patients have increased serum acrolein levels and impaired neutrophil functions. These results suggest that acrolein suppresses neutrophil function and contributes to the switch in the neutrophil phenotype in glioma.