Background: Depression is a mental disorder that poses a serious threat to human health. Adult hippocampal neurogenesis (AHN) is closely associated with the efficacy of antidepressants. Chronic treatment with corticosterone (CORT), a well-validated pharmacological stressor, induces depressive-like behaviors and suppresses AHN in experimental animals. However, the possible mechanisms of chronic CORT action remain elusive.

Mounting attention has been focused on defects in macroautophagy/autophagy and the autophagy-lysosomal pathway (ALP) in cerebral ischemia. TFEB (transcription factor EB)-mediated induction of ALP has been recently considered as the common mechanism in ameliorating the pathological lesion of myocardial ischemia and neurodegenerative diseases. Here we explored the vital role of TFEB in permanent middle cerebral artery occlusion (pMCAO)-mediated dysfunction of ALP and ischemic insult in rats. The results showed that ALP function was first enhanced in the early stage of the ischemic process, especially in neurons of the cortex, and this was accompanied by increased TFEB expression and translocation to the nucleus, which was mediated at least in part through activation by PPP3/calcineurin. At the later stages of ischemia, a gradual decrease in the level of nuclear TFEB was coupled with a progressive decline in lysosomal activity, accumulation of autophagosomes and autophagy substrates, and exacerbation of the ischemic injury. Notably, neuron-specific overexpression of TFEB significantly enhanced ALP function and rescued the ischemic damage, starting as early as 6 h and even lasting to 48 h after ischemia. Furthermore, neuron-specific knockdown of TFEB markedly reversed the activation of ALP and further aggravated the neurological deficits and ischemic outcome at the early stage of pMCAO. These results highlight neuronal-targeted TFEB as one of the key players in the pMCAO-mediated dysfunction of ALP and ischemic injury, and identify TFEB as a promising target for therapies aimed at neuroprotection in cerebral ischemia. Abbreviations: AAV, adeno-associated virus; AIF1/IBA1, allograft inflammatory factor 1; ALP, autophagy-lysosomal pathway; CQ, chloroquine; CTSB, cathepsin B; CTSD, cathepsin D; CsA, cyclosporin A; GFAP, glial fibrillary acidic protein; LAMP, lysosomal-associated membrane protein; LC3, microtubule-associated protein 1 light chain 3; MAP2, microtubule-associated protein 2; mNSS, modified Neurological Severity Score; MTOR, mechanistic target of rapamycin kinase; OGD, oxygen and glucose deprivation; pMCAO, permanent middle cerebral artery occlusion; RBFOX3/NeuN, RNA binding fox-1 homolog 3; SQSTM1, sequestosome1; TFEB, transcription factor EB; TTC, 2,3,5-triphenyltetrazolium chloride.

Ischemic stroke is a cerebrovascular disease with high morbidity, high mortality, and high disability, representing a serious threat to human life and health. Clinically, the extensive injury caused by ischemic stroke results from ischemia-reperfusion (I/R) injury thrombolytic treatment. However, there are few reports on the use of medications in the subacute stage of cerebral I/R. Baicalein (5,6,7-trihydroxyflavone) is a biologically active ingredient extracted from the root of Scutellaria baicalensis Georgi. In the present study, we investigated the therapeutic effect of baicalein administered in the subacute phase of cerebral I/R injury in a rat model of ischemia induced by occlusion of the middle cerebral artery (MCA). Rats were treated daily with baicalein (200 mg/kg, i.g.) in the subacute phase (24 h after reperfusion) for 7 days. The results showed that baicalein significantly reduced neurobehavioral deficits and decreased brain infarct volume from 18.99% to 7.41%. Immunofluorescence analysis of the ischemic penumbra showed that baicalein significantly reduced expression of the M1 marker, cluster of differentiation (CD) 16 and CD86, and increased expression of the M2 marker, CD 163 and CD206, indicating that baicalein inhibited M1 transformation and promoted M2 transformation of microglia/macrophage to inhibit neuroinflammation. Moreover, baicalein suppressed NF-κB signaling by reducing IκBα phosphorylation and nuclear translocation of NF-κB/p65, which decreased the release of the pro-inflammatory factors IL-6, IL-18, and TNF-α. In addition, baicalein reduced phosphorylation of JNK, ERK and p38, which are involved modulation of microglia/macrophage M1/M2 polarization. Western blot analysis of apoptosis- and autophagy-related proteins showed that baicalein increased the Bcl-2/Bax ratio and reduced caspase-3 expression to decrease neuronal apoptosis and ameliorate neuronal loss. Baicalein also decreased the LC3-II/LC3-I ratio and promoted phosphorylation of the PI3K/Akt/mTOR signaling pathway which implied inhibition of autophagy. These observations suggest that baicalein exerts neuroprotective effects by reducing neuroinflammation, apoptosis and autophagy, and protects against cerebral I/R injury in the subacute phase in vivo.