Sham Control Research Articles

(276) Light Emitting Diodes (LED) as a Potential Therapy for Erectile Function: A Preclinical Study in a Cavernous Nerve Injury Model

Abstract Introduction Erectile dysfunction (ED) induced by nerve injury including prostatectomy poorly responds to PDE5 inhibitor and requires invasive treatment. Light emitting diodes (LED) in some spectral range, is able to irradiate cells without overheating for its low energy consumption and to enhance mitochondrial metabolism, intracellular energy transfer, and thereby cell proliferation and tissue regeneration in some organs. We, herein, investigated whether and how LED therapy may improve penile erection in mouse cavernous nerve injury (CNI) - induced ED model Objective To identify the optimal LED parameters using molecular, histological, and erectile function analysis in cavernous nerve injury. Methods Eight week old male mice (n=120) were divided into 5 age-matched groups, a sham operation group receiving heat treatment, a CNI group receiving heat treatment, a CNI group receiving red light of LED (wavelength; 660 ±3% nm), a CNI group receiving near infrared light of LED (wavelength; 830 ±2% nm), and a CNI group receiving combination of both LED light. Mice were exposed to LED light for 30 minutes on the ventral side including penis for 5 consecutive days after CNI operation. Molecular study, histopathological analysis of the penis and erectile function test were done 2 weeks following the treatment in each subject. Results CNI group with heat treatment had much lower maximum and total ICP than sham control group. In contrast, three groups with LED light irradiation exhibited considerably improved maximum and total ICP, and the group with both LED treatments had the best erectile performance, reaching 90% of the sham surgery group. Histologically, mice treated with RED, NIR, or LED irradiation had more cavernous pericytes and endothelial cells than CNI mice after sham operation. In addition, all three CNI groups with various LED lengths improved neuroprotection and brain regeneration in CNI mice. All groups treated with LED showed increased expression of βIII-tubulin, neurofilament-1, and nNOS in cavernous tissue and dorsal nerve bundle. Additionally, LED treatment in three groups, particularly the combination group, significantly increased βIII-tubulin, S100, and myelin basic protein levels, indicating axonal integrity or expansion in the dorsal root and major pelvic ganglions. Molecular analysis of downstream signals of neural regeneration or angiogenesis showed that 3 groups with LED treatment increased phosphorylated PI3K, neurotrophic factors like BDNF, NGF, and NT-3, and angiogenic factors like angiopoietin1 and vascular endothelial growth factor in cavernous tissue and llpopolysaccharide-treated PC-12 cells. The TUNEL assay and BrdU staining of these tissue and cells showed lower cell apotosis and proliferation in LED-treated groups. Conclusions The results show that LED therapy with RED, NIR, preferably combination of each wavelength improves penile erection by neural progection or regeneration in cavernous nerve injury induced-ED. This noninvasive therapy may be useful in near future for ED refractory to PDE5 inhibitor. As far as we are aware, this is the first research to demonstrate the advantage of LED therapy, low level in sexual dyfunction field. Disclosure No.

Does vibrotactile stimulation of the auricular vagus nerve enhance working memory? A behavioral and physiological investigation

BackgroundWorking memory is essential to a wide range of cognitive functions and activities. Transcutaneous auricular vagus nerve stimulation (taVNS) is a promising method to improve working memory performance. However, the feasibility and scalability of electrical stimulation are constrained by several limitations, such as auricular discomfort and inconsistent electrical contact. ObjectiveWe aimed to develop a novel and practical method, vibrotactile taVNS, to improve working memory. Further, we investigated its effects on arousal, measured by skin conductance and pupil diameter. MethodThis study included 20 healthy participants. Behavioral response, skin conductance, and eye tracking data were concurrently recorded while the participants performed N-back tasks under three conditions: vibrotactile taVNS delivered to the cymba concha, earlobe (sham control), and no stimulation (baseline control). ResultsIn 4-back tasks, which demand maximal working memory capacity, active vibrotactile taVNS significantly improved the performance metric d′ compared to the baseline but not to the sham. Moreover, we found that the reduction rate of d′ with increasing task difficulty was significantly smaller during vibrotactile taVNS sessions than in both baseline and sham conditions. Arousal, measured as skin conductance and pupil diameter, declined over the course of the tasks. Vibrotactile taVNS rescued this arousal decline, leading to arousal levels corresponding to optimal working memory levels. Moreover, pupil diameter and skin conductance level were higher during high-cognitive-load tasks when vibrotactile taVNS was delivered to the concha compared to baseline and sham. ConclusionOur findings suggest that vibrotactile taVNS modulates the arousal pathway and could be a potential intervention for enhancing working memory.

Antioxidant and hepatoprotective activities of Juniperus excelsa M. Bieb against bile duct ligation-induced cholestasis.

Cholestasis is caused by a malfunction of the biliary liver system. Oxidative stress plays an essential role in the progression of cholestasis. This study aimed to investigate the antioxidant and hepatoprotective effects of ethanolic extract of Juniperus excelsa M. Bieb (JE) fruits on hepatic impairment induced by bile duct ligation (BDL) in rats. Forty male Wistar rats were randomly divided into 4 groups; sham control + vehicle (SC), BDL + vehicle (BDL), BDL + JE extract (BDL + JE), and SC + extract (SC + JE). One day after surgery, the animals were treated with vehicle or ethanolic extract of JE (500 mg/kg/day) for 7 days. Finally, the blood was taken for biochemical and oxidative stress analysis. Furthermore, the liver tissue of rats was removed for histological examination. Treatment with the extract of JE decreased the ALP level, whereas it enhanced total protein content compared to the BDL group. Also, JE increased the activity of SOD and GPx, as well as FRAP content compared to the BDL group; while it did not significantly affect the levels of MDA and inflammation markers. However, JE could not improve BDL-induced histopathological alterations in hepatic tissue. This study demonstrated that JE may be useful as an adjuvant therapy by attenuating ALP activity, increasing serum total protein and FRAP content, as well as improving the antioxidant enzymes activity of SOD and GPx. However, further research is warranted to explore the other underlying mechanisms of action.

Motor Recovery after Spinal Cord Trauma and Effect of Local Hypothermia in a Porcine Experimental Model

Abstract This study was aimed to assess the therapeutic potential (expressed by improvement of pelvic extremities motor functions) of a 5-hour local application of hypothermia with cold saline (4 °C), or saline at room temperature (≈ 24 °C) conveyed via perfusion chamber placed epidurally over the epicenter of spinal cord lesion in minipigs paraplegic due to acute spinal cord injuries (SCIs) inflicted through L3 laminectomy with the force of 8N, 15N, or 18N by a computer operated contusion apparatus. Eighteen 5–8-month-old minipigs (Göttingen-Minnesota-Liběchov crossbreed strains weighing 28–35 kg) were randomly divided into 6 subgroups (each containing three animals) another 3 minipigs were added as sham controls. To evaluate the pelvic extremities motor recovery was used the porcine 20-point neurological scale. Regular evaluations of motor scores showed gradual spontaneous recovery of this parameter in all experimental animals, however, the best results achieved minipigs after SCI inflicted by 8N impacts. The data achieved in the study suggest that local application of therapeutic hypothermia (TH) is well tolerated and may improve functional outcomes after SCI. Further experimental and preclinical studies in different SCI animal models are required before the introduction of the method in healthcare practice.

Possible therapeutic effect of combined platelet-rich plasma and stem cells on induced knee osteoarthritis in adult male albino rat: histological and immunohistochemical study

Osteoarthritis (OA) is a degenerative joint disease that results from breakdown of joint cartilage and underlying bone. It is the most common form of arthritis among elderly individuals. The present study aimed to investigate the effect of intra‑articular injection of platelet-rich plasma (PRP) and bone marrow mesenchymal stem cells (BMSCs), both in combination and separately in the treatment of monoiodoacetate (MIA) induced OA in rats. Seventy adult male albino rats were randomly divided into seven groups, 10 rats in each (control, sham control, OA-induced, PRP, BMSCs, PRP+BMSCs and recovery). Histological examination was done using haematoxylin and eosin, alcian blue and Masson’s trichrome stains. Immunohistochemical staining for collagen type II antigen and proliferating cell nuclear antigen (PCNA) was carried out. Measurements of serum interleukin 6 (IL-6) was performed using ELISA. Expression of ACAN gene was done using real-time polymerase chain reaction (PCR) and histomorphometric measurements were analyzed. Radiographs were acquired from the knee joint. Light microscopic examination of articular cartilage (AC) from OA-induced group showed disorganization of chondrocytes, loss of zonation, pyknotic nuclei, empty lacunae and absent tidemark. There was a significant decrease in AC thickness, chondrocytes count, optical density of collagen and proteoglycan content, area percent of collagen II in the matrix in OA group compared to control and sham groups. These degenerative histomorphological alterations were associated with significant increase in serum IL-6 and decrease in ACAN gene expression. Radiographs showed narrowing of joint space and subchondral sclerosis. Intraarticular injection of PRP, BMSCs and PRP+BMSCs improved the previously mentioned alterations. Concomitant administration of PRP+BMSCs showed better effect than using PRP and BMSCs alone. In conclusion, PRP potentiated the effects of BMSCs on the repair of MIA-induced AC damage in rats. Injection of combination of PRP with BMSCs could be considered as a promising therapy for OA in patients.

Targeted delivery of miR-1 to the heart using clinical contrast ultrasound

Pathological left-ventricle hypertrophy is a cardiovascular disorder resulting in the thickening of the ventricle wall due to abnormal cardiomyocyte growth. The downregulation of miR-1 in hypertrophic cardiomyocytes has been identified as an early disease marker, with the delivery of miR-1 as a promising treatment strategy. Ultrasound and microbubbles offer an exciting approach to image-guided and site-specific cardiac gene delivery. The objective of this study is to show the feasibility of viable ultrasound and microbubble-mediated delivery of miR-1 in vivo. Sprague–Dawley rats were injected via tail vein with a suspension of miR-1 mimic (0.6 mg/kg) and Definity. The rats were then treated with a high MI (1.34) flash sequence for 20 min using a C5-2 probe with a Phillips iU22. Delivery was confirmed on isolated heart tissue with RT-qPCR and Western blots for miR-1 and protein expression, respectively. miR-1 delivery in healthy male rats resulted in a 1.33-fold (p = 0.05) increase in miR-1 as compared to sham controls. This resulted in a decrease in hypertrophic protein expression (1.18-fold in TWF1, p = 0.17; 1.23-fold in MEF2A, p = 0.07; 1.25-fold in CX43, p = 0.03). Our data demonstrate the feasibility of using ultrasound and microbubbles as an image-guided delivery method for molecular therapeutics in cardiovascular disorders.

Comparative evaluation of two different xenogenic acellular matrices on full-thickness skin wound healing.

The purpose of the study was to compare the healing potential of bubaline small intestinal matrix (bSIM) and fish swim bladder matrix (FSBM) on full-thickness skin wounds in rabbits. Four full-thickness skin wounds (each 20×20mm) were created on the dorsum of 18 rabbits that were divided into three groups based on treatment: untreated sham control (I), implanted with double layers of bSIM (II) and implanted with double layers of FSBM (III). Macroscopic, immunologic and histologic observations were made to evaluate wound healing. Gross healing progression in the bSIM and FSBM groups showed significantly (p<0.05) less wound contraction compared with the sham group. The IgG concentration in rabbit sera was significantly (p<0.05) lower in the FSBM group compared with the bSIM group by enzyme-linked immunosorbent assay. The stimulation index of peripheral blood lymphocytes was significantly (p<0.05) lower in the FSBM group compared with the bSIM group by the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Implantation of FSBM resulted in improved re-epithelialisation, neovascularisation and fibroplasia. The FSBM is a more effective dermal substitute when compared with the bSIM for full-thickness skin wound repair in rabbit.

SPAK inhibitor ZT-1a attenuates reactive astrogliosis and oligodendrocyte degeneration in a mouse model of vascular dementia.

Astrogliosis and white matter lesions (WML) are key characteristics of vascular contributions to cognitive impairment and dementia (VCID). However, the molecular mechanisms underlying VCID remain poorly understood. Stimulation of Na-K-Cl cotransport 1 (NKCC1) and its upstream kinases WNK (with no lysine) and SPAK (the STE20/SPS1-related proline/alanine-rich kinase) play a role in astrocytic intracellular Na+ overload, hypertrophy, and swelling. Therefore, in this study, we assessed the effect of SPAK inhibitor ZT-1a on pathogenesis and cognitive function in a mouse model of VCID induced by bilateral carotid artery stenosis (BCAS). Following sham or BCAS surgery, mice were randomly assigned to receive either vehicle (DMSO) or SPAK inhibitor ZT-1a treatment regimen (days 14-35 post-surgery). Mice were then evaluated for cognitive functions by Morris water maze, WML by exvivo MRI-DTI analysis, and astrogliosis/demyelination by immunofluorescence and immunoblotting. Compared to sham control mice, BCAS-Veh mice exhibited chronic cerebral hypoperfusion and memory impairments, accompanied by significant MRI DTI-detected WML and oligodendrocyte (OL) death. Increased activation of WNK-SPAK-NKCC1-signaling proteins was detected in white matter tissues and in C3d+ GFAP+ cytotoxic astrocytes but not in S100A10+ GFAP+ homeostatic astrocytes in BCAS-Veh mice. In contrast, ZT-1a-treated BCAS mice displayed reduced expression and phosphorylation of NKCC1, decreased astrogliosis, OL death, and WML, along with improved memory functions. BCAS-induced upregulation of WNK-SPAK-NKCC1 signaling contributes to white matter-reactive astrogliosis, OL death, and memory impairment. Pharmacological inhibition of the SPAK activity has therapeutic potential for alleviating pathogenesis and memory impairment in VCID.

ISSLS PRIZE in Basic Science 2024: superiority of nucleus pulposus cell- versus mesenchymal stromal cell-derived extracellular vesicles in attenuating disc degeneration and alleviating pain

PurposeTo investigate the therapeutic potential of extracellular vesicles (EVs) derived from human nucleus pulposus cells (NPCs), with a specific emphasis on Tie2-enhanced NPCs, compared to EVs derived from human bone marrow-derived mesenchymal stromal cells (BM-MSCs) in a coccygeal intervertebral disc degeneration (IDD) rat model.MethodsEVs were isolated from healthy human NPCs cultured under standard (NPCSTD-EVs) and Tie2-enhancing (NPCTie2+-EVs) conditions. EVs were characterized, and their potential was assessed in vitro on degenerative NPCs in terms of cell proliferation and senescence, with or without 10 ng/mL interleukin (IL)-1β. Thereafter, 16 Sprague–Dawley rats underwent annular puncture of three contiguous coccygeal discs to develop IDD. Phosphate-buffered saline, NPCSTD-EVs, NPCTie2+-EVs, or BM-MSC-derived EVs were injected into injured discs, and animals were followed for 12 weeks until sacrifice. Behavioral tests, radiographic disc height index (DHI) measurements, evaluation of pain biomarkers, and histological analyses were performed to assess the outcomes of injected EVs.ResultsNPC-derived EVs exhibited the typical exosomal morphology and were efficiently internalized by degenerative NPCs, enhancing cell proliferation, and reducing senescence. In vivo, a single injection of NPC-derived EVs preserved DHI, attenuated degenerative changes, and notably reduced mechanical hypersensitivity. MSC-derived EVs showed marginal improvements over sham controls across all measured outcomes.ConclusionOur results underscore the regenerative potential of young NPC-derived EVs, particularly NPCTie2+-EVs, surpassing MSC-derived counterparts. These findings raise questions about the validity of MSCs as both EV sources and cellular therapeutics against IDD. The study emphasizes the critical influence of cell type, source, and culture conditions in EV-based therapeutics.

Therapeutic implication of MicroRNA-320a antagonist in attenuating blood clots formed during venous thrombosis.

Venous thrombosis (VT) is a complex multi-factorial disease and a major health concern worldwide. Its clinical implications include deep vein thrombosis (DVT) and pulmonary embolism (PE). VT pathogenesis involves intricate interplay of various coagulants and anti-coagulants. Growing evidences from epidemiological studies have shown that many non-coding microRNAs play significant regulatory role in VT pathogenesis by modulating expressions of large number of gene involved in blood coagulation. Present study aimed to investigate the effect of human micro RNA (hsa-miR)-320a antagonist on thrombus formation in VT. Surgery was performed on Sprague-Dawley (SD) rats, wherein the inferior vena cava (IVC) was ligated to introduce DVT. Animals were divided into four groups (n = 5 in each group); Sham controls (Sham), IVC ligated-DVT (DVT), IVC ligated-DVT + transfection reagent (DVT-NC) and IVC ligated-DVT + miR320a antagonist (DVT-miR-320a antagonist). IVC was dissected after 6h and 24h of surgery to estimate thrombus weight and coagulatory parameters such as levels of D-dimer, clotting time and bleeding time. Also, ELISA based biochemical assays were formed to assess toxicity of miRNA antagonist in animals. Our experimental analysis demonstrated that there was a marked reduction in size of thrombus in hsa-miR-320a antagonist treated animals, both at 6h and 24h. There was a marked reduction in D-dimer levels in hsa-miR-320a antagonist treated animals. Also, blood clotting time was delayed and bleeding time was increased significantly in hsa-miR-320a antagonist treated rats compared to the non-treated and Sham rats. There was no sign of toxicity in treated group compared to control animals. Hsa-miR-320a antagonist could be promising therapeutic target for management of VT.

Natural intraepithelial lymphocyte populations rise during necrotic enteritis in chickens.

Intraepithelial lymphocytes (IEL) reside in the epithelium at the interface between the contents of the intestinal lumen and the sterile environment of the lamina propria. Because of this strategic location, IEL play a crucial role in various immunological processes, ranging from pathogen control to tissue stability. In mice and humans, IEL exhibit high diversity, categorized into induced IEL (conventional CD4 and CD8αβ T cells) and natural IEL (TCRαβCD8αα, TCRγδ, and TCRneg IEL). In chickens, however, the subpopulations of IEL and their functions in enteric diseases remain unclear. Thus, we conducted this study to investigate the role of IEL populations during necrotic enteritis (NE) in chickens. At 14 days of age, sixty-three Specific-pathogen-free (SPF) birds were randomly assigned to three treatments: Control (sham challenge), Eimeria maxima challenge (EM), and Eimeria maxima + Clostridium Perfringens (C. Perfringens) co-challenge (EM/CP). The EM and EM/CP birds were infected with Eimeria maxima at day 14 of age, and EM/CP birds were additionally orally inoculated with C. perfringens at days 18 and 19 of age. Birds were weighed at days 18, 20, and 26 of age to assess body weight gain (BWG). At 20 days of age (1 day-post C. perfringens infection; dpi), and 26 days of age (7 dpi), 7 birds per treatment were euthanized, and jejunum was harvested for gross lesion scores, IEL isolation, and gene expression. The EM/CP birds exhibited subclinical NE disease, lower BWG and shorter colon length. The Most changes in the IEL populations were observed at 1 dpi. The EM/CP group showed substantial increases in the total number of natural IEL subsets, including TCRαβ+CD4-CD8-, TCRαβ+CD8αα+, TCRγδ+, TCRneg and innate CD8α (iCD8α) cells by at least two-fold. However, by 7 dpi, only the number of TCRαβ+CD4-CD8- and TCRαβ+CD8αα+ IEL maintained their increase in the EM/CP group. The EM/CP group had significantly higher expression of proinflammatory cytokines (IL-1β and IFN-γ) and Osteopontin (OPN) in the jejunum at 1 dpi. These findings suggest that natural IEL with innate and innate-like functions might play a critical role in the host response during subclinical NE, potentially conferring protection against C. perfringens infection.

Integrative Analysis of Cytokine and Lipidomics Datasets Following Mild Traumatic Brain Injury in Rats.

Traumatic brain injury (TBI) is a significant source of disability in the United States and around the world and may lead to long-lasting cognitive deficits and a decreased quality of life for patients across injury severities. Following the primary injury phase, TBI is characterized by complex secondary cascades that involve altered homeostasis and metabolism, faulty signaling, neuroinflammation, and lipid dysfunction. The objectives of the present study were to (1) assess potential correlations between lipidome and cytokine changes after closed-head mild TBI (mTBI), and (2) examine the reproducibility of our acute lipidomic profiles following TBI. Cortices from 54 Sprague Dawley male and female rats were analyzed by ultra-high-performance liquid chromatography mass spectrometry (LC-MS) in both positive and negative ionization modes and multiplex cytokine analysis after single (smTBI) or repetitive (rmTBI) closed-head impacts, or sham conditions. Tissue age was a variable, given that two cohorts (n = 26 and n = 28) were initially run a year-and-a-half apart, creating inter-batch variations. We annotated the lipidome datasets using an in-house data dictionary based on exact masses of precursor and fragment ions and removed features with statistically significant differences between sham control batches. Our results indicate that lipids with high-fold change between injury groups moderately correlate with the cytokines eotaxin, IP-10, and TNF-α. Additionally, we show a significant decrease in the pro-inflammatory markers IL-1β and IP-10, TNF-α, and RANTES in the rmTBI samples relative to the sham control. We discuss the major challenges in correlating high dimensional lipidomic data with functional cytokine profiles and the implications for understanding the biological significance of two related but disparate analysis modes in the study of TBI, an inherently heterogeneous neurological disorder.

Investigation of the effects of in ovo taurine injection on hatching characteristics and stress reduction potential.

In ovo application is the process of administering some nutrients or components into the egg. The main purpose of this application is to ensure that some nutrients are provided to chicks with a short incubation period. Few studies were conducted with taurine in fertile eggs; especially, no observation of hatchability and chick quality has been found. In addition, taurine has an anti-stress impact that fights oxidative factors. To assess the hatchability and chick quality after in ovo taurine administration. To determine the stress that may occur as a result of in ovo application and whether taurine has a stress-reducing effect. A total of 1200 fertile eggs from a 34-week-old broiler breeder (Ross 308) flock were categorized into 4 groups with 75 eggs per replicate: control (uninjected), taurine group (0.30mL dissolved taurine in distilled water), sham control (sterile distilled water) and perforation (eggs perforated and then waxed). On day 14 of incubation, an in ovo injection was administered to the albumen. Data on hatching parameters and hepatic HSP70 levels were obtained using relevant formulas and western blotting, respectively. Control chicks exhibited higher hatchability than other groups, with the taurine group showing the lowest hatchability. The HSP70 levels were the highest in the perforation group compared to the control group. An increase of 21.37% in the taurine group and 83.45% in the sham control group was observed compared to the control group. The findings suggest that in ovo application may induce increased stress, whereas taurine may have positive effects in mitigating the stress caused by in ovo application.

The priority of mesenchymal stem cells treated with melatonin in ameliorating thyroid impairment induced by amiodarone. Thorough look on the apoptotic pathway

Amiodarone treatment has been associated with thyroid alterations. This work was planned to consider therapeutic outcome of MSCs versus MSCs treated with melatonin in minimizing amiodarone –induced deviations in thyroid.We handed-down 50 male Wistar rats, then distributed them into 5 groups; I, II, III, IV, V; control, sham control, amiodarone treated, amiodarone and MSCs treated, and amiodarone, MSCs and melatonin treated groups respectively. Light microscopic examination; levels of T3, T4 and TSH, Oxidative/antioxidative tissue markers, immune-histochemical staining (Bcl2, BAX, iNOS) and real time PCR (IL-6 and VEGF and Caspase 3) were done. Results of group III showed degenerated follicles, decreased follicular cell count and diminished colloid. Some of the follicles were dilated with signs of inflammatory response and apoptosis. Increased collagen deposition in group III was marked. The positive immune-reactive cells of Bcl-2 was decreased and that of BAX and iNOS was increased, also T3 and T4 levels were significantly decreased, but TSH was significantly increased in group III comparing it to the group I. There were highly significant diminution in both SOD and GPx and upsurge in MDA intensities in groups III, IV when correlated to the control. In group IV and V the aforementioned values were restored. The PCR results showed significant increase in IL-6 and VEGF and Caspase 3 in group III compared to the control one, whereas, their values in groups IV and V were reestablished. It is concluded that stem cells can to a great extent ameliorate the thyroid damage induced by amiodarone.But, Adding melatonin to the stem cells culture was found to have auxiliary beneficial effect in the improving the thyroid structure and function.

Frontal midline theta transcranial alternating current stimulation enhances early consolidation of episodic memory

Evidence implicating theta rhythms in declarative memory encoding and retrieval, together with the notion that both retrieval and consolidation involve memory reinstatement or replay, suggests that post-learning theta rhythm modulation can promote early consolidation of newly formed memories. Building on earlier work employing theta neurofeedback, we examined whether theta-frequency transcranial alternating stimulation (tACS) can engender effective consolidation of newly formed episodic memories, compared with beta frequency stimulation or sham control conditions. We compared midline frontal and posterior parietal theta stimulation montages and examined whether benefits to memory of theta upregulation are attributable to consolidation rather than to retrieval processes by using a washout period to eliminate tACS after-effects between stimulation and memory assessment. Four groups of participants viewed object pictures followed by a free recall test during three study-test cycles. They then engaged in tACS (frontal theta montage/parietal theta montage/frontal beta montage/sham) for a period of 20 min, followed by a 2-h break. Free recall assessments were conducted after the break, 24 h later, and 7 days later. Frontal midline theta-tACS induced significant off-line retrieval gains at all assessment time points relative to all other conditions. This indicates that theta upregulation provides optimal conditions for the consolidation of episodic memory, independent of mental-state strategies.

Knockdown of ANGPTL2 promotes left ventricular systolic dysfunction by upregulation of NOX4 in mice.

Background: Angiopoietin-like 2 (ANGPTL2) is a pro-inflammatory and pro-oxidant circulating protein that predicts and promotes chronic inflammatory diseases such as atherosclerosis in humans. Transgenic murine models demonstrated the deleterious role of ANGPTL2 in vascular diseases, while deletion of ANGPTL2 was protective. The nature of its role in cardiac tissues is, however, less clear. Indeed, in adult mice knocked down (KD) for ANGPTL2, we recently reported a mild left ventricular (LV) dysfunction originating from a congenital aortic valve stenosis, demonstrating that ANGPTL2 is essential to cardiac development and function. Hypothesis: Because we originally demonstrated that the KD of ANGPTL2 protected vascular endothelial function via an upregulation of arterial NOX4, promoting the beneficial production of dilatory H2O2, we tested the hypothesis that increased cardiac NOX4 could negatively affect cardiac redox and remodeling and contribute to LV dysfunction observed in adult Angptl2-KD mice. Methods and results: Cardiac expression and activity of NOX4 were higher in KD mice, promoting higher levels of cardiac H2O2 when compared to wild-type (WT) mice. Immunofluorescence showed that ANGPTL2 and NOX4 were co-expressed in cardiac cells from WT mice and both proteins co-immunoprecipitated in HEK293 cells, suggesting that ANGPTL2 and NOX4 physically interact. Pressure overload induced by transverse aortic constriction surgery (TAC) promoted LV systolic dysfunction in WT mice but did not further exacerbate the dysfunction in KD mice. Importantly, the severity of LV systolic dysfunction in KD mice (TAC and control SHAM) correlated with cardiac Nox4 expression. Injection of an adeno-associated virus (AAV9) delivering shRNA targeting cardiac Nox4 expression fully reversed LV systolic dysfunction in KD-SHAM mice, demonstrating the causal role of NOX4 in cardiac dysfunction in KD mice. Targeting cardiac Nox4 expression in KD mice also induced an antioxidant response characterized by increased expression of NRF2/KEAP1 and catalase. Conclusion: Together, these data reveal that the absence of ANGPTL2 induces an upregulation of cardiac NOX4 that contributes to oxidative stress and LV dysfunction. By interacting and repressing cardiac NOX4, ANGPTL2 could play a new beneficial role in the maintenance of cardiac redox homeostasis and function.

Placebo response to sham electroacupuncture in patients with chronic functional constipation: A secondary analysis.

Chronic severe functional constipation is a common disease that requires novel and effective treatment strategies. Acupuncture might constitute a promising therapeutic approach for chronic constipation, but it reportedly engenders an enhanced placebo response. Herein, we aimed to determine the magnitude of the placebo responses to electroacupuncture for constipation and identify its influencing factors. In this secondary analysis of a multicenter randomized trial, patients were randomized to a sham electroacupuncture group, which was administered for 8 weeks in 24 sessions of superficial needling at nonacupoints. The placebo response rates were assessed using three responder criteria: ≥3 complete spontaneous bowel movements per week (CSBMs/week), overall CSBM, and sustained CSBM. Logistic regression with backward selection method was employed to identify the potential factors that affected the placebo response. Overall, 539 patients were included in the study. The placebo response rate was 10.58%, 9.46%, and 9.09% according to the three aforementioned criteria, respectively. Those who exhibited more CSBMs/week at baseline were more likely to respond to sham electroacupuncture. Among patients with more than one CSBM per week at baseline, the response rates for achieving the criteria of ≥3 CSBMs per week, overall CSBM, and sustained CSBM were 25.40%, 30.16%, and 22.22%. Electroacupuncture did not demonstrate a large placebo response in functional constipation treatment. The number of CSBMs at baseline may be a moderator of the placebo response to sham electroacupuncture. Thus, superficial needling sham acupuncture may be considered an ideal sham control for clinical trials of chronic constipation.

Effect of His Bundle Pacing on Abnormal Myocardial Fatty Acid and Glucose Metabolism Induced by Right Ventricular Pacing.

Metabolic disorder is noted for pacing-induced cardiomyopathy. The benefits of His bundle pacing over right ventricular (RV) pacing in preventing pacing-induced cardiomyopathy from a metabolic perspective are yet to be fully understood. Three pig groups were established for this study: sham control, RV pacing (RV pacing for 6 months), and His pacing (RV pacing for 6 months, followed by His bundle pacing for 3 months). Complete atrioventricular block was created in the last 2 groups. Left ventricular function and dyssynchrony were assessed via echocardiography, while proteins linked to metabolism, endoplasmic reticulum stress, and inflammation in left ventricular myocardium were examined. The RV pacing group had significantly more left ventricular mechanical dyssynchrony compared with the other groups. The RV pacing group exhibited triglyceride and diacylglycerol accumulation in cardiomyocytes and higher expression of binding immunoglobulin protein and tumor necrosis factor-α than the other groups. Additionally, the expression of CD36 was activated, while the expression of hormone-sensitive lipase was downregulated in the RV pacing group compared with the His pacing and sham control groups. Furthermore, the expressions of GLUT4 and pyruvate dehydrogenase were higher in the RV pacing group than the sham control and His pacing groups. Notably, the abnormal fatty acid and glucose metabolic pathways in the left ventricular myocardium during RV pacing could be corrected by His bundle pacing. His bundle pacing can mitigate the abnormal metabolism disorders, endoplasmic reticulum stress, and inflammation induced during RV pacing and may contribute to the superiority of conduction system pacing over RV pacing in reducing heart failure hospitalization.

Integrated multi-omic analysis identifies fatty acid binding protein 4 as a biomarker and therapeutic target of ischemia–reperfusion injury in steatotic liver transplantation

Background and aimsDue to a lack of donor grafts, steatotic livers are used more often for liver transplantation (LT). However, steatotic donor livers are more sensitive to ischemia–reperfusion (IR) injury and have a worse prognosis after LT. Efforts to optimize steatotic liver grafts by identifying injury targets and interventions have become a hot issue.MethodsMouse LT models were established, and 4D label-free proteome sequencing was performed for four groups: normal control (NC) SHAM, high-fat (HF) SHAM, NC LT, and HF LT to screen molecular targets for aggravating liver injury in steatotic LT. Expression detection of molecular targets was performed based on liver specimens from 110 donors to verify its impact on the overall survival of recipients. Pharmacological intervention using small-molecule inhibitors on an injury-related target was used to evaluate the therapeutic effect. Transcriptomics and metabolomics were performed to explore the regulatory network and further integrated bioinformatics analysis and multiplex immunofluorescence were adopted to assess the regulation of pathways and organelles.ResultsHF LT group represented worse liver function compared with NC LT group, including more apoptotic hepatocytes (P < 0.01) and higher serum transaminase (P < 0.05). Proteomic results revealed that the mitochondrial membrane, endocytosis, and oxidative phosphorylation pathways were upregulated in HF LT group. Fatty acid binding protein 4 (FABP4) was identified as a hypoxia-inducible protein (fold change > 2 and P < 0.05) that sensitized mice to IR injury in steatotic LT. The overall survival of recipients using liver grafts with high expression of FABP4 was significantly worse than low expression of FABP4 (68.5 vs. 87.3%, P < 0.05). Adoption of FABP4 inhibitor could protect the steatotic liver from IR injury during transplantation, including reducing hepatocyte apoptosis, reducing serum transaminase (P < 0.05), and alleviating oxidative stress damage (P < 0.01). According to integrated transcriptomics and metabolomics analysis, cAMP signaling pathway was enriched following FABP4 inhibitor use. The activation of cAMP signaling pathway was validated. Microscopy and immunofluorescence staining results suggested that FABP4 inhibitors could regulate mitochondrial membrane homeostasis in steatotic LT.ConclusionsFABP4 was identified as a hypoxia-inducible protein that sensitized steatotic liver grafts to IR injury. The FABP4 inhibitor, BMS-309403, could activate of cAMP signaling pathway thereby modulating mitochondrial membrane homeostasis, reducing oxidative stress injury in steatotic donors.Graphical

A nine-month repeat-dose intravaginal ring (Ovaprene) irritation study in sheep

ObjectivesOvaprene is a novel, investigational, intravaginal hormone-free monthly ring contraceptive designed for use in women of reproductive age to be worn over multiple weeks (one menstrual cycle). The objective of this work was to evaluate the safety of Ovaprene during a nine-month repeat-dose sheep study. Study designIn addition to traditional safety endpoints such as histopathological evaluation of the sheep female reproductive tract, vaginal fluids were collected and tested for released iron over time. Also, the amount of iron in the rings was assessed following removal, and serum iron levels were measured. There were four sheep in each group (Ovaprene group and sham group). ResultsThere were no macroscopic clinical findings. There was minimal to mild, mixed or mononuclear cell infiltration present in all levels of vagina (cranial, mid, and caudal) from all animals including sham controls based on post-study necropsy. The female reproductive tract from animals treated with the Ovaprene ring was comparable to the sham controls. The concentrations of serum iron in sheep treated with Ovaprene ring were similar compared to a sham treated animal. The average amount of ferrous gluconate released from Ovaprene over the 29-day period of use was 175 mg of the approximately 512 mg nominally loaded into the rings. ConclusionsOverall, the Ovaprene devices were well-tolerated in female sheep. ImplicationsThis study should support a chronic (e.g., one year) contraceptive efficacy study in women.