The positive role of transforming growth factor-ß1 in ischemic stroke.

Ischemic stroke is one of the most disabling and fatal diseases around the world. The damaged brain tissues will undergo excessive autophagy, vascular endothelial cells injury, blood-brain barrier (BBB) impairment and neuroinflammation after ischemic stroke. However, there is no unified viewpoint on the underlying mechanism of brain damage. Transforming growth factor-ß1 (TGF-ß1), as a multi-functional cytokine, plays a crucial role in the intricate pathological processes and helps maintain the physiological homeostasis of brain tissues through various signaling pathways after ischemic stroke. In this review, we summarize the protective role of TGF-ß1 in autophagic flux, BBB, vascular remodeling, neuroinflammation and other aspects after ischemic stroke. Based on the review, we believe that TGF-ß1 could serve as a key target for treating ischemic stroke.

Evaluation of left ventricular function in hypertensive athletic patients with different circadian rhythms using stratified strain technique

Purpose:With the stratified strain approach, the changes in the longitudinal mechanical condition of the left ventricular myocardium in hypertensive athleticpatients with a normal left ventricular ejection fraction (LVEF) have been evaluated. Materials and Methods:Based on circadian rhythm, 48 dippers and 53 non-dippers are selected among 103 hypertensionathleticpatients with normal LVEF (LVEF > 50%). The stratified strain approach is used to measure the longitudinal strain (GLS) of the subendocardial, middle,and epicardial myocardium during left ventricular systole. The relationship between the stratified strain and the circadian rhythm in hypertensive individuals has been analysed. Results:show that the levels of IVST, LVPWT, and LVMI in the non-dipper group are considerably higher than those in the dipper group, while there is not statistically significant between the two groups in terms of LVDd, LVDs, LA, or EF. Myocardial strain analysis reveals that the left ventricular long axis strain (GLS) in the non-dipper group is considerably greater than that in the dipper group, And the difference is statistically significant (P< 0.05). But the GLSendo between the non-dipper group and the dipper group did not vary significantly (P<0.05). In GLSMid, GLSepi, there is no significant difference, and the difference is not statistically significant(P>0.05).Conclusion：Ultrasonic stratified strain technique can quantitatively measure and analyze the longitudinal stratified strain of the myocardium in each segment of the left ventricle in hypertensive athleticpatients and evaluate their early left ventricular mechanics changes, providing a better diagnostic method for early clinical determination of myocardial involvement in athleticpatients. (AU)

[Pathogenic mechanisms of impaired neuronal autophagy flux after ischemic stroke].

Autophagy is a metabolic process in which damaged organelles, obsolete proteins, excess cytoplasmic components, and even pathogens are presented to lysosomes for degradation via autophagosomes. It includes 4 processes: the initiation of autophagy, the formation of autophagosomes, the fusion of autophagosomes with lysosomes, and the degradation and removal of autophagic substrates within autophagic lysosomes. When these processes are continuous, it is called autophagy flux. Blockage of one or certain steps in the autophagy/lysosome signaling pathway can lead to impaired autophagy flux. Numerous studies have shown that impaired autophagy flux is an important cause of neuronal damage in the ischemic penumbra after stroke. This paper summarized research progress in the pathological mechanisms that cause impaired neuronal autophagy flux after ischemic stroke and discusses methods to improve neuronal autophagy flux, in order to provide a reference for an in-depth investigation of the pathological injury mechanisms after stroke.

Berberine Alleviates Ischemic Brain Injury by Enhancing Autophagic Flux via Facilitation of TFEB Nuclear Translocation.

Berberine has been demonstrated to alleviate cerebral ischemia/reperfusion injury, but its neuroprotective mechanism has yet to be understood. Studies have indicated that ischemic neuronal damage was frequently driven by autophagic/lysosomal dysfunction, which could be restored by boosting transcription factor EB (TFEB) nuclear translocation. Therefore, this study investigated the pharmacological effects of berberine on TFEB-regulated autophagic/lysosomal signaling in neurons after cerebral stroke. A rat model of ischemic stroke and a neuronal ischemia model in HT22 cells were prepared using middle cerebral artery occlusion (MCAO) and oxygen-glucose deprivation (OGD), respectively. Berberine was pre-administered at a dose of 100[Formula: see text]mg/kg/d for three days in rats and 90[Formula: see text][Formula: see text]M in HT22 neurons for 12[Formula: see text]h. 24[Formula: see text]h after MCAO and 2[Formula: see text]h after OGD, the penumbral tissues and OGD neurons were obtained to detect nuclear and cytoplasmic TFEB, and the key proteins in the autophagic/lysosomal pathway were examined using western blot and immunofluorescence, respectively. Meanwhile, neuron survival, infarct volume, and neurological deficits were assessed to evaluate the therapeutic efficacy. The results showed that berberine prominently facilitated TFEB nuclear translocation, as indicated by increased nuclear expression in penumbral neurons as well as in OGD HT22 cells. Consequently, both autophagic activity and lysosomal capacity were simultaneously augmented to alleviate the ischemic injury. However, berberine-conferred neuroprotection could be greatly counteracted by lysosomal inhibitor Bafilomycin A1 (Baf-A1). Meanwhile, autophagy inhibitor 3-Methyladenine (3-MA) also slightly neutralized the pharmacological effect of berberine on ameliorating autophagic/lysosomal dysfunction. Our study suggests that berberine-induced neuroprotection against ischemic stroke is elicited by enhancing autophagic flux via facilitation of TFEB nuclear translocation in neurons.

Psychophysiological restorative potential in cancer patients by virtual reality (VR)-based perception of natural environment.

The positive significance of nature to human' self-reported well-being has been widely confirmed, but less attention has been paid to the study of cancer patients, as well as the role of time on the restorative effects. Therefore, using virtual reality (VR) and the inclusion of patients with esophageal and gastrointestinal cancer as participants, this study conducted indoor experiments to explore patients' psychophysiological recovery through the perception of five different environmental types with three to five interventions per week. There were 63 participants selected from the People's Hospital in Shaanxi Province. Depending on their psychophysiological state, they would participate in three to five interventions in a week to compare the number of interventions needed to achieve maximum restoration. The five environmental types utilized varied in land cover, vegetation structure, and landscape characteristics, and were identified as blue space (BS), open green space (OGS), semi-open green space (SOS), closed green space (CGS), and gray space (GrS). Before and after viewing landscapes, the changes of psychophysiological indicators were measured to explore the influence of different environmental types on participants. The results showed that the participants preferred and received the highest perceived restorative potentials in BS and lastly, GrS. The green and blue spaces measurably increased positive emotions and perceived restoration while a decreasing negative emotions and the heart rate (HR) compared with the GrS. Participants had the highest level of relaxation while their eyes were closed in the EEG baseline stage. Moreover, participants received the most relaxation when they contacted with nature three times a week, which indicated that excessive natural participation may not be conducive to the sustained development of cancer patients' psychophysiological health. Instead of field appreciation, VR could be utilized to increase the access of cancer patients to nature and then be used as an approach to landscape interaction.

[Activation of JNK induces apoptosis to autophagy conversion and enhances the survival of oxygen-glycogen deprived rat neurons].

Objective To investigate the efficacy and mechanism of c-Jun N-terminal kinase (JNK) in boosting survival of oxygen glucose deprivation (OGD) rat neurons. Methods The cortex neurons from fetal rats were primarily cultured to prepare a model of OGD neurons in vitro, and the characteristic endpoints were filtered to intervene with JNK inducer anisomycin (AN), respectively. The cells were randomly divided into control group, solvent control group (a same volume of solvent DMSO was added into the culture medium of the OGD neuron), AN group (OGD neurons were treated with JNK inducer AN for 5 hours at the end of OGD). After that, Western blotting and immunofluorescence cytometry were respectively performed to detect the protein expressions in OGD neurons, including beclin 1, microtubule-associated protein 1 light chain 3 (LC3), B cell lymphoma 2 (Bcl2), caspase-3, P62, ubiquitin, cathepsin B and lysosomal associated membrane protein 1 (LAMP1). The cell activity was evaluated by CCK-8 assay, and the axon length was measured by IPP software. Results Activation of JNK significantly promoted the expressions of beclin 1, LC3, and Bcl2, and markedly reduced the content of beclin 1-Bcl2 complex and attenuated the expressions of P62 and ubiquitin. Meanwhile, the expressions of cathepsin B and LAMP1 were not obviously altered. In this way, the survival rate of OGD neurons was improved. Conclusion Activation of JNK exerts a neuroprotective effect by facilitating dissociation of beclin 1-Bcl2 and inducing a switch from apoptosis to autophagy in OGD neurons.

Melibiose Confers a Neuroprotection against Cerebral Ischemia/Reperfusion Injury by Ameliorating Autophagy Flux via Facilitation of TFEB Nuclear Translocation in Neurons.

Autophagic/lysosomal dysfunction is a critical pathogenesis of neuronal injury after ischemic stroke. Trehalose has been validated to restore the impaired autophagy flux by boosting transcription factor EB (TFEB) nuclear translocation, but orally administrated trehalose can be greatly digested by intestinal trehalase before entering into brain. Melibiose (MEL), an analogue of trehalose, may thoroughly exert its pharmacological effects through oral administration due to absence of intestinal melibiase. The present study was to investigate whether melibiose could also confer a neuroprotection by the similar pharmacological mechanism as trehalose did after ischemic stroke. The rats were pretreated with melibiose for 7 days before middle cerebral artery occlusion (MCAO) surgery. Twenty-four hours following MCAO/reperfusion, the cytoplasmic and nuclear TFEB, and the proteins in autophagic/lysosomal pathway at the penumbra were detected by western blot and immunofluorescence, respectively. Meanwhile, the neurological deficit, neuron survival, and infarct volume were assessed to evaluate the therapeutic outcomes. The results showed that the neurological injury was significantly mitigated in MCAO+MEL group, compared with that in MCAO group. Meanwhile, nuclear TFEB expression in neurons at the penumbra was significantly promoted by melibiose. Moreover, melibiose treatment markedly enhanced autophagy flux, as reflected by the reinforced lysosomal capacity and reduced autophagic substrates. Furthermore, the melibiose-elicited neuroprotection was prominently counteracted by lysosomal inhibitor Bafilomycin A1 (Baf-A1). Contrarily, reinforcement of lysosomal capacity with EN6 further improved the neurological performance upon melibiose treatment. Our data suggests that melibiose-augmented neuroprotection may be achieved by ameliorating autophagy flux via facilitation of TFEB nuclear translocation in neurons after ischemic stroke.

Autophagy Elicits Neuroprotection at the Subacute Phase of Transient Cerebral Ischaemia but Has Few Effects on Neurological Outcomes After Permanent Ischaemic Stroke in Rats.

OBJECTIVE: Autophagy was prominently activated by cerebral ischaemia. This study was to investigate the exact role of autophagy in ischaemic stroke. METHODS: Two rat models of transient middle cerebral artery occlusion (tMCAO) and permanent MCAO (pMCAO) were prepared. The brain tissues in the penumbra were obtained to observe the dynamic variations of autophagy activity with Beclin1 and LC3 antibodies by Western blotting. At the characteristic time points, when autophagy activity was markedly elevated or reduced, the autophagy activation signaling was intervened with rapamycin and 3-methyladenine, respectively. Thereafter, key proteins in the autopahgic/lysosomal pathway were detected with the antibodies of LC3, p62, ubiquitin, LAMP-1 and cathepsin B. Meanwhile, TTC staining, neurological score and immunofluorescence were performed to evaluate brain infarct volume, neurological deficit and neuron survival, respectively. RESULTS: Both Beclin1 and LC3 expression levels were remarkably altered at 6 h, 12 h, 2 days and 7 days after tMCAO. Interestingly, the dynamic changes of autophagy activity following pMCAO were identical to those after tMCAO. Neither autophagy induction nor autophagy inhibition was able to ameliorate the pMCAO-induced neurological injury due to lysosomal dysfunction, as indicated by low levels of LAMP-1 and cathepsin B, accompanied with the accumulation of LC3-II, ubiquitin and insoluble p62. Comparatively, autophagy induction elicited overt neuroprotection at 2 and 7 days after tMCAO, and this neuroprotection might be elicited by the enhancement of autophagy flux. CONCLUSION: Our study suggests that autophagy confers neuroprotection at the subacute phase of tMCAO but has few effects on neurological outcomes after pMCAO.

Enriched environment boosts the post-stroke recovery of neurological function by promoting autophagy.

Autophagy is crucial for maintaining cellular homeostasis, and can be activated after ischemic stroke. It also participates in nerve injury and repair. The purpose of this study was to investigate whether an enriched environment has neuroprotective effects through affecting autophagy. A Sprague-Dawley rat model of transient ischemic stroke was prepared by occlusion of the middle cerebral artery followed by reperfusion. One week after surgery, these rats were raised in either a standard environment or an enriched environment for 4 successive weeks. The enriched environment increased Beclin-1 expression and the LC3-II/LC3-I ratio in the autophagy/lysosomal pathway in the penumbra of middle cerebral artery-occluded rats. Enriched environment-induced elevations in autophagic activity were mainly observed in neurons. Enriched environment treatment also promoted the fusion of autophagosomes with lysosomes, enhanced the lysosomal activities of lysosomal-associated membrane protein 1, cathepsin B, and cathepsin D, and reduced the expression of ubiquitin and p62. After 4 weeks of enriched environment treatment, neurological deficits and neuronal death caused by middle cerebral artery occlusion/reperfusion were significantly alleviated, and infarct volume was significantly reduced. These findings suggest that neuronal autophagy is likely the neuroprotective mechanism by which an enriched environment promotes recovery from ischemic stroke. This study was approved by the Animal Ethics Committee of the Kunming University of Science and Technology, China (approval No. 5301002013855) on March 1, 2019.

Neuronal autophagy aggravates microglial inflammatory injury by downregulating CX3CL1/fractalkine after ischemic stroke.

Ischemic stroke often induces excessive neuronal autophagy, resulting in brain damage; meanwhile, inflammatory responses stimulated by ischemia exacerbate neural injury. However, interactions between neuronal autophagy and microglial inflammation following ischemic stroke are poorly understood. CX3CL1/fractalkine, a membrane-bound chemokine expressed on neurons, can suppress microglial inflammation by binding to its receptor CX3CR1 on microglia. In the present study, to investigate whether autophagy could alter CX3CL1 expression on neurons and consequently change microglial inflammatory activity, middle cerebral artery occlusion (MCAO) was established in Sprague-Dawley rats to model ischemic stroke, and tissues from the ischemic penumbra were obtained to evaluate autophagy level and microglial inflammatory activity. MCAO rats were administered 3-methyladenine (autophagy inhibitor) or Tat-Beclin 1 (autophagy inducer). Western blot assays were conducted to quantify expression of Beclin-1, nuclear factor kappa B p65 (NF-κB), light chain 3B (LC3B), and CX3CL1 in ischemic penumbra. Moreover, immunofluorescence staining was performed to quantify numbers of LC3B-, CX3CL1-, and Iba-1-positive cells in ischemic penumbra. In addition, enzyme linked immunosorbent assays were utilized to analyze concentrations of tumor necrosis factor alpha (TNF-α), interleukin 6 (IL-6), interleukin 1 beta (IL-1ß), and prostaglandin E2 (PGE2). A dry/wet weight method was used to detect brain water content, while 2,3,5,-triphenyltetrazolium chloride staining was utilized to measure infarct volume. The results demonstrated that autophagy signaling (Beclin-1 and LC3B expression) in penumbra was prominently activated by MCAO, while CX3CL1 expression on autophagic neurons was significantly reduced and microglial inflammation was markedly activated. However, after inhibition of autophagy signaling with 3-methyladenine, CX3CL1 expression on neurons was obviously increased, whereas Iba-1 and NF-κB expression was downregulated; TNF-α, IL-6, IL-1ß, and PGE2 levels were decreased; and cerebral edema was obviously mitigated. In contrast, after treatment with the autophagy inducer Tat-Beclin 1, CX3CL1 expression on neurons was further reduced; Iba-1 and NF-κB expression was increased; TNF-α, IL-6, IL-1ß, and PGE2 levels were enhanced; and cerebral edema was aggravated. Our study suggests that ischemia-induced neuronal autophagy facilitates microglial inflammatory injury after ischemic stroke, and the efficacy of this process may be associated with downregulated CX3CL1 expression on autophagic neurons.

Differential variations of autophagy and apoptosis in permanent focal cerebral ischaemia rat model.

OBJECTIVE: Autophagy and apoptosis coexist in stroke, but the relationship between effects and complex is poorly understood. Herein, we investigated dynamic changes of autophagy and apoptosis at the penumbra in permanent cerebral ischaemia. METHODS: Sprague-Dawley rat models were prepared by middle cerebral artery occlusion. The autophagy and apoptosis were evaluated by Western blotting and immunofluorescence with LC3-II and cleaved caspase-3, respectively. The neurological deficit score and infarct volume were assessed. RESULTS: The results showed that the expressions of LC3-II and cleaved caspase-3 were gradually increased from 1 to 5 hours, and reached maximum at 5 hours after stroke. After that, LC3-II expression was significantly declined, but cleaved caspase-3 expression was only mildly reduced from 6 hours to 3 days. Surprisingly, at 4 days after stroke, the autophagy was abruptly increased again, but the apoptosis was considerably and continuously decreased. The severity of the neurological deficit was in accordance with the increase of infarct expansion. CONCLUSIONS: Our results showed that autophagy and apoptosis were simultaneously activated within 12 hours after stroke. Four days later, LC3-II expression was significantly increased, while cleaved caspase-3 expression was considerably decreased, implying that there might be a transition from apoptosis to autophagy at the subacute phase of stroke.

Dynamic changes in neuronal autophagy and apoptosis in the ischemic penumbra following permanent ischemic stroke.

The temporal dynamics of neuronal autophagy and apoptosis in the ischemic penumbra following stroke remains unclear. Therefore, in this study, we investigated the dynamic changes in autophagy and apoptosis in the penumbra to provide insight into potential therapeutic targets for stroke. An adult Sprague-Dawley rat model of permanent ischemic stroke was prepared by middle cerebral artery occlusion. Neuronal autophagy and apoptosis in the penumbra post-ischemia were evaluated by western blot assay and immunofluorescence staining with antibodies against LC3-II and cleaved caspase-3, respectively. Levels of both LC3-II and cleaved caspase-3 in the penumbra gradually increased within 5 hours post-ischemia. Thereafter, levels of both proteins declined, especially LC3-II. The cerebral infarct volume increased slowly 1-4 hours after ischemia, but subsequently increased rapidly until 5 hours after ischemia. The severity of the neurological deficit was positively correlated with infarct volume. LC3-II and cleaved caspase-3 levels were high in the penumbra within 5 hours after ischemia, and after that, levels of these proteins decreased at different rates. LC3-II levels were reduced to a very low level, but cleaved caspase-3 levels remained high 72 hours after ischemia. These results indicate that there are temporal differences in the activation status of the autophagic and apoptotic pathways. This suggests that therapeutic targeting of these pathways should take into consideration their unique temporal dynamics.

Hydrothermal synthesis and characterization of aluminum-free Mn-ß zeolite: a catalyst for phenol hydroxylation.

Zeolite beta, especially heteroatomic zeolite beta, has been widely used in the industries of fine chemicals and petroleum refining because of its outstanding thermal stability, acid resistance, and unique 3-D open-frame structure. In this paper, aluminum-free Mn-ß zeolite was hydrothermally synthesized in the SiO2-MnO2-(TEA)2O-NaF-H2O system. The effect of the chemical composition of the precursor mixture to the crystallization of the Al-free Mn-ß zeolite was investigated. The synthesized Al-free Mn-ß zeolite was characterized by inductively coupled plasma (ICP), XRD, thermogravimetric/differential thermal analysis (TG/DTA), N2 adsorption-desorption, FT-IR, UV-vis, X-ray photoelectron spectroscopy (XPS), and scanning electron microscope (SEM). The results show that the synthesized zeolite has a structure of ß zeolite with good crystallinity and Mn ions present in the framework of the zeolite. The synthesized Al-free Mn-ß zeolite shows great catalytic activity toward the phenol hydroxylation reaction using H2O2 as the oxidant. Approximately 35% of phenol conversion and ∼98% of dihydroxybenzene selectivity can be obtained under the optimal conditions.

[Analysis of survival and prognostic factors in extremely preterm infants].

OBJECTIVE: To investigate the survival rate, complications during hospitalization, and prognostic factors in extremely preterm infants (gestational age less than 28 weeks) in the neonatal intensive care unit (NICU). METHODS: A retrospective analysis was performed on 90 extremely preterm infants who were admitted to the NICU between January 2011 and March 2013 to investigate the perinatal data, delivery and resuscitation, ventilation/oxygen supply during hospitalization, mortality, and the incidence of severe (grade III/IV) intraventricular hemorrhage (IVH), bronchopulmonary dysplasia (BPD), retinopathy of prematurity (ROP), patent ductus arteriosus (PDA), and septicemia. RESULTS: Among the 90 extremely preterm infants, the gestational age, birth weight, overall survival rate, mortality, and withdrawal rate were 26±1 weeks, 898±165 g, 57%, 9%, and 34%, respectively. The incidence rates of neonatal respiratory distress syndrome, BPD, PDA, ROP, and grade III/IV IVH were 88%, 85%, 69%, 68%, and 31%, respectively. The surviving infants had a mean length of hospital stay of 83±18 days and a mean weight at discharge of 2 419±300 g. The multivariate logistic regression analysis showed that grade III/IV IVH and pulmonary hemorrhage were high-risk factors for death or withdrawal, while antenatal corticosteroids were the protective factor for outcome (P<0.05). CONCLUSIONS: The survival rate of extremely preterm infants is still much lower than that in developed countries. grade III/IV IVH, and pulmonary hemorrhage are important prognostic factors.

Evaluation of protective efficacy conferred by a recombinant Mycobacterium bovis BCG expressing a fusion protein of Ag85A-ESAT-6.

BACKGROUND: We previously constructed a recombinant bacille Calmette-Guérin (rBCG-AE) strain that could express a fused Ag85A-ESAT-6 protein. That study suggested that the rBCG-AE strain was able to induce a higher titer of antibody and elicit a more long-lived and stronger Th1-type cellular immune responses than the parental BCG strain, the rBCG-A strain (i.e., expressing Ag85A), or the rBCG-E strain (i.e., expressing ESAT-6). METHODS: In the current study, we further investigated the strain's protective efficacy against Mycobacterium tuberculosis H37Rv infection in BALB/c mice through evaluating organ bacterial loads, lung histopathology, lung immunohistochemistry, and net weight gain or loss by using conventional BCG, rBCG-A, and rBCG-E as the controls. RESULTS: From the 3rd to 9th weeks after the challenge infection, the bacterial counts were significantly lower in tissues (e.g., spleen and lung tissues) in the mice immunized with rBCG-AE than in the control group, but were higher than the counts in the BCG group. The pathological damage in the lung tissues of the rBCG-AE group gradually improved from the 6th to 9th weeks after being infected with M. tuberculosis H37Rv, but the score of pathological changes in the rBCG-AE group was obviously higher than the score in the BCG group. There was no difference in the percentage of IFN-γ and iNOS positive cells in the lung tissues of the rBCG-AE and BCG groups. CONCLUSION: The results suggest that rBCG-AE can not promote protective efficacy against M. tuberculosis H37Rv infection, compared to the BCG vaccine.

[Study on immunogenicity elicited by a recombinant vaccine of rBCG-Rv3133c to fight against dormancy Mycobacterium tuberculosis].

To obtain a vaccine to defend from dormancy Mycobacterium tuberculosis, we constructed the recombinant Bacilli Calmette-Guérin (BCG) vaccine with Rv3133c encoding dormancy-correlated transcriptional regulatory protein DosR in Mycobacterium tuberculosis as a target gene, and evaluated its immunogenicity in BALB/c mice. In this study, we constructed the recombinant plasmids of rpMV361-Rv3133c using gene colon technology. We then transformed BCG strains with above-mentioned plasmids to obtain recombinant vaccine of rBCG-Rv3133c. We used the rBCG strains successfully constructed to vaccinate in BALB/c mice. 30d and 180d after immunization, the specific antibody titers were determined to investigate humoral responses induced by recombinant vaccine. We detected changes of splenocyte subsets of CD4+T, CD8+ T cells and cytokine of IFN-gamma secreted by splenocytes for evaluation of cellular immune responses. The results showed that the rBCG-Rv3133c was able to induce higher levels of antibody titer, stronger proliferative responses and higher IFN-gamma production comparing with BCG vaccine. The results also suggested that this recombinant vaccine was a more efficacious tuberculosis vaccine for further study.