μM Propofol Research Articles

In Vitro Effect of Propofol on the Expression of Genes Involved in Inflammation and Apoptosis in Corneal Activated Keratocytes.

We investigated the effect of propofol (0.5, 5, and 50 μM) on the gene expression of inflammatory cytokines [ IL-1β , IL-6 , transforming growth factor β ( TGF-β ), and LIF ] and apoptosis process ( BCL-2 and Bax ) in corneal activated keratocytes (CAKs). CAKs (10 6 cells/10 cm 2 ) were exposed to propofol at a concentration of 0.5, 5, and 50 μM for 24 hours at 37°C. The control group did not receive propofol at the same time or under the same condition. Ribonucleic acid (RNA) extraction, complementary DNA (cDNA) synthesis, and real-time polymerase chain reaction (PCR) were performed to quantify the relative expression of IL-1β , IL-6 , TGF-β , LIF , BCL-2 , and Bax expression in the treated versus control cells. The results of this study showed that propofol treatment (0.5 and 5 μM) led to the downregulation of IL-1β and IL-6 gene expression in CAKs. TGF-β (with a role in fibrogenesis) was not changed in 0.5 and 5 μM propofol-treated CAKs, whereas CAKs treated with 50 μM propofol showed upregulation of the TGF-β gene. LIF (with a role in regeneration) was upregulated in 0.5 and 5 μM propofol-treated CAKs. The BCL-2/Bax ratio (as the antiapoptosis index) was increased in CAKs treated with 0.5 μM propofol and indicated the induction of an antiapoptotic effect. We showed that CAKs treatment with propofol, at concentrations of 0.5 and 5 μM, could decrease the expression of genes related to inflammation and enhance the genes associated with cell regeneration. While 50 μM propofol treatment might induce CAK fibrogenesis. This proof-of-concept study could preserve a groundwork for future drug design for the treatment of corneal stromal diseases and ocular regenerative medicine.

The effect of propofol on chemosensitivity of paclitaxel in cervical cancer cells.

Propofol is a drug with potential anticancer effect. This study aimed to explore the effect of propofol on chemosensitivity of cervical cancer cells to paclitaxel. HeLa and CaSki cells were selected for drug experiments. Cell viability was evaluated via CCK-8 assay, and the combination index (CI) was calculated by CompuSyn software. A clinically relevant concentration and IC30 of propofol were selected in combination with 5 nM paclitaxel. BrdU incorporation, transwell, and flow cytometry assays were utilized to evaluate cell proliferation, migration, invasion, and apoptosis. The expression of β-tubulin, stathmin 1, and GAPDH proteins was evaluated by Western blot. The stathmin 1 cDNA plasmid was used to establish stathmin 1-overexpressing CaSki cells. At clinically relevant concentrations (0-80 μM), propofol did not affect cancer cell viability, but high concentrations (100-800 μM) reduced cell viability. The CI values of propofol with IC30 (200 μM in HeLa; 400 μM in CaSki) combined with 5 nM paclitaxel were <1. The effect of propofol with IC30 combined with paclitaxel on cell proliferation, migration, invasion, and apoptosis were stronger than individual effect, while 30 μM propofol had no effect. The Western blot results showed 30 μM propofol did not affect β-tubulin and stathmin 1 expression in cells, although paclitaxel upregulated β-tubulin expression while downregulating stathmin 1 expression. Compared with paclitaxel alone, cotreatment with propofol at its IC30 and paclitaxel decreased stathmin 1 expression but had no effect on β-tubulin expression. High stathmin 1 expression weakened the effect of paclitaxel on cell viability and apoptosis, while propofol partially reversed these effect. Propofol at clinically relevant concentrations had no effect on the malignant biological behaviors of cervical cancer cells, while propofol at high concentrations decreased.Propofol with IC30 and paclitaxel had synergetic effect on cancer cells through a reduction in stathmin 1 expression.

The Mechanism of TNF-α-Mediated Accumulation of Phosphorylated Tau Protein and Its Modulation by Propofol in Primary Mouse Hippocampal Neurons: Role of Mitophagy, NLRP3, and p62/Keap1/Nrf2 Pathway.

Background Neuroinflammation-induced phosphorylated Tau (p-Tau) deposition in central nervous system contributes to neurodegenerative disorders. Propofol possesses neuroprotective properties. We investigated its impacts on tumor necrosis factor-α (TNF-α)-mediated p-Tau deposition in neurons. Methods Mouse hippocampal neurons were exposed to propofol followed by TNF-α. Cell viability, p-Tau, mitophagy, reactive oxygen species (ROS), NOD-like receptor protein 3 (NLRP3), antioxidant enzymes, and p62/Keap1/Nrf2 pathway were investigated. Results TNF-α promoted p-Tau accumulation in a concentration- and time-dependent manner. TNF-α (20 ng/mL, 4 h) inhibited mitophagy while increased ROS accumulation and NLRP3 activation. It also induced glycogen synthase kinase-3β (GSK3β) while inhibited protein phosphatase 2A (PP2A) phosphorylation. All these effects were attenuated by 25 μM propofol. In addition, TNF-α-induced p-Tau accumulation was attenuated by ROS scavenger, NLRP3 inhibitor, GSK3β inhibitor, or PP2A activator. Besides, compared with control neurons, 100 μM propofol decreased p-Tau accumulation. It also decreased ROS and NLRP3 activation, modulated GSK3β/PP2A phosphorylation, leaving mitophagy unchanged. Further, 100 μM propofol induced p62 expression, reduced Keap1 expression, triggered the nuclear translocation of Nrf2, and upregulated superoxide dismutase (SOD) and heme oxygenase-1 (HO-1) expression, which was abolished by p62 knockdown, Keap1 overexpression, or Nrf2 inhibitor. Consistently, the inhibitory effect of 100 μM propofol on ROS and p-Tau accumulation was mitigated by p62 knockdown, Keap1 overexpression, or Nrf2 inhibitor. Conclusions In hippocampal neurons, TNF-α inhibited mitophagy, caused oxidative stress and NLRP3 activation, leading to GSK3β/PP2A-dependent Tau phosphorylation. Propofol may reduce p-Tau accumulation by reversing mitophagy and oxidative stress-related events. Besides, propofol may reduce p-Tau accumulation by modulating SOD and HO-1 expression through p62/Keap1/Nrf2 pathway.

The effect of propofol on hypoxia- and TNF-α-mediated BDNF/TrkB pathway dysregulation in primary rat hippocampal neurons.

AimsHypoxia and inflammation may lead to BDNF/TrkB dysregulation and neurological disorders. Propofol is an anesthetic with neuroprotective properties. We wondered whether and how propofol affected BDNF/TrkB pathway in hippocampal neurons and astrocytes.MethodsPrimary rat hippocampal neurons and astrocytes were cultured and exposed to propofol followed by hypoxia or TNF‐α treatment. The expression of BDNF and the expression/truncation/phosphorylation of TrkB were measured. The underlying mechanisms were investigated.ResultsHypoxia and TNF‐α reduced the expression of BDNF, which was reversed by pretreatment of 25 μM propofol in hippocampal neurons. Furthermore, hypoxia and TNF‐α increased the phosphorylation of ERK and phosphorylation of CREB at Ser142, while reduced the phosphorylation of CREB at Ser133, which were all reversed by 25 μM propofol and 10 μM ERK inhibitor. In addition, hypoxia or TNF‐α did not affect TrkB expression, truncation, or phosphorylation in hippocampal neurons and astrocytes. However, in hippocampal neurons, 50 μM propofol induced TrkB phosphorylation, which may be mediated by p35 expression and Cdk5 activation, as suggested by the data showing that blockade of p35 or Cdk5 expression mitigated propofol‐induced TrkB phosphorylation.ConclusionsPropofol modulated BDNF/TrkB pathway in hippocampal neurons via ERK/CREB and p35/Cdk5 under the condition of hypoxia or TNF‐α exposure.

Post-treatment with propofol inhibits inflammatory response in LPS-induced alveolar type II epithelial cells.

Over-inflammation and severe lung injury are major causes of morbidity and mortality in patients with coronavirus disease 2019 (COVID-19). With the COVID-19 pandemic, an increasing number of patients with preexisting lung injury and inflammation are undergoing surgery or artificial ventilation under sedation in intensive care units, where 2,6-diisopropylphenol (propofol) is a commonly used drug for sedation. The aim of the present study was to investigate whether post-inflammation treatment with propofol protects epithelial type II cells against inflammation in an in vitro model of inflammation. The A549 cell line, characterised as epithelial type II cells, were exposed to lipopolysaccharide (LPS) for 2 h and subsequently treated with different concentrations of propofol (0, 10, 25 or 50 µM) for 3 h. Western blot and reverse transcription-quantitative PCR analyses were used to detect the protein and mRNA expression levels, respectively, of CD14 and Toll-like receptor 4 (TLR4). Immunofluorescence staining was used to detect the in situ CD14 and TLR4 expression in epithelial type II cells. Tumor necrosis factor (TNF)-α production was also examined using ELISA. LPS significantly increased the expression of CD14 and TLR4, as well as the secretion of TNF-α. Post-treatment with 25 and 50 µM propofol of the LPS-treated cells significantly decreased CD14 and TLR4 expression, as well as TNF-α secretion, compared with the cells treated with LPS only, indicating that post-treatment with propofol alleviated inflammation and this effect was dose-dependent. The present study suggested that treatment with propofol after LPS administration has a protective effect on epithelial type II cells.

Propofol via Antioxidant Property Attenuated Hypoxia-Mediated Mitochondrial Dynamic Imbalance and Malfunction in Primary Rat Hippocampal Neurons.

Background Hypoxia may induce mitochondrial abnormality, which is associated with a variety of clinical phenotypes in the central nervous system. Propofol is an anesthetic agent with neuroprotective property. We examined whether and how propofol protected hypoxia-induced mitochondrial abnormality in neurons. Methods Primary rat hippocampal neurons were exposed to propofol followed by hypoxia treatment. Neuron viability, mitochondrial morphology, mitochondrial permeability transition pore (mPTP) opening, mitochondrial membrane potential (MMP), and adenosine triphosphate (ATP) production were measured. Mechanisms including reactive oxygen species (ROS), extracellular regulated protein kinase (ERK), protein kinase A (PKA), HIF-1α, Drp1, Fis1, Mfn1, Mfn2, and Opa1 were investigated. Results Hypoxia increased intracellular ROS production and induced mPTP opening, while reducing ATP production, MMP values, and neuron viability. Hypoxia impaired mitochondrial dynamic balance by increasing mitochondrial fragmentation. Further, hypoxia induced the translocation of HIF-1α and increased the expression of Drp1, while having no effect on Fis1 expression. In addition, hypoxia induced the phosphorylation of ERK and Drp1ser616, while reducing the phosphorylation of PKA and Drp1ser637. Importantly, we demonstrated all these effects were attenuated by pretreatment of neurons with 50 μM propofol, antioxidant α-tocopherol, and ROS scavenger ebselen. Besides, hypoxia, propofol, α-tocopherol, or ebselen had no effect on the expression of Mfn1, Mfn2, and Opa1. Conclusions In rat hippocampal neurons, hypoxia induced oxidative stress, caused mitochondrial dynamic imbalance and malfunction, and reduced neuron viability. Propofol protected mitochondrial abnormality and neuron viability via antioxidant property, and the molecular mechanisms involved HIF-1α-mediated Drp1 expression and ERK/PKA-mediated Drp1 phosphorylation.

Effects of Dexmedetomidine on the Expression of Cytochrome C Oxidase in Neonatal Mouse Primary Hippocampal Neurons Cultured in Propofol

To observe the effects of propofol and dexmedetomidine on the expression of cytochrome c oxidase in primary hippocampal neurons isolated from neonatal mice. C57 neonatal mice were sacrificed within 24 h of birth and hippocampal tissue was collected. Primary hippocampal neurons were cultured in vitro and then randomly divided into a control group (0 μM propofol and 0 μM dexmedetomidine), propofol group (20 μM propofol and 0 μM dexmedetomidine) and propofol+dexmedetomidine group (20 μM propofol and 10 μM dexmedetomidine). The neurons were incubated for 1 h. Neuron morphology was observed under a light microscope. The expression of cytochrome c protein was detected semiquantitatively by Western blot and immunofluorescence staining. Compared with those in the control group, the primary hippocampal neurons in the propofol group were morphologically degenerated, with sparse axons and dendrites; the expression of cytochrome c protein was significantly higher in the propofol group (p<0.0001). The morphology of primary hippocampal neurons in the propofol+dexmedetomidine group was similar to that of primary hippocampal neurons in the control group, with plump and bright cell bodies and thickened neurites intertwined in a neuron network; additionally, cytochrome c protein expression was significantly lower in the hippocampal neurons in the propofol+dexmedetomidine group than in the hippocampal neurons in propofol group (p<0.0001). Dexmedetomidine reversed propofol induced injury and morphological abnormalities of primary hippocampal neurons. The mechanism may be related to the downregulation of cytochrome c protein expression.

Effect of Propofol and Etomidate on the Proliferation, Cell-cycle Distribution, Apoptosis and Necrosis of Pancreatic Tumour Cells.

The influence of surgical interventions and anaesthesiological procedures on tumour progression was investigated as early as the 1920s. In current cancer management, the perioperative phase is increasingly being considered a vulnerable period with an increased risk of tumour cell dissemination due to medication, surgical manipulation, and immunosuppression. The extent to which narcotics administered in the perioperative setting influence the oncological outcomes of patients with pancreatic cancer is still unclear. To investigate the effect of propofol and etomidate on the proliferation, cell-cycle distribution, apoptosis, and necrosis of pancreatic tumour cells in vitro, PaTu 8988t and Panc-1 pancreatic cancer cells were treated with 0-1,000 μM propofol or etomidate for 24 h each. Cell proliferation was measured with enzyme-linked immunosorbent-bromodeoxyuridine assay. The apoptosis rate was analysed with annexin V staining and the cell-cycle distribution with flow cytometry. Propofol at 1,000 μM induced apoptosis and inhibited cell proliferation. The cell cycle showed an increased S-phase and reduced cells in the G1-phase. At 100 μM, propofol significantly inhibited proliferation of the pancreatic cancer cell line PaTu 8988t and reduced cells in the G2-phase in the cell cycle. Etomidate had no effects on cell-cycle distribution, proliferation, apoptosis, and necrosis at the concentrations used. In this study, propofol was shown to have anticancer effects by induction of apoptosis and inhibition of cell proliferation, while etomidate did not affect pancreatic cancer cells. However, it is too early to make any recommendation for changes in clinical practice and further clinical studies are warranted to investigate the effect of anaesthetics on cancer progression.

Propofol Ameliorates ox-LDL-Induced Endothelial Damage Through Enhancing Autophagy via PI3K/Akt/m-TOR Pathway: A Novel Therapeutic Strategy in Atherosclerosis.

Objective: Atherosclerosis (AS) represents a common age-associated disease, which may be accelerated by oxidized low-density lipoprotein (ox-LDL)-induced endothelial cell injury. This study aimed to investigate the effects of Propofol on ox-LDL-induced endothelial damage and the underlying molecular mechanisms. Methods: Human umbilical vein endothelial cells (HUVECs) were exposed to ox-LDL to induce endothelial damage. HUVECs were pretreated with 0, 5, 25 and 100°μM Propofol, followed by exposure to 100°μg/ml ox-LDL for 24°h. Cell viability was assessed by cell counting kit-8 (CCK-8) assay. The expression of autophagy- and apoptosis-related proteins was detected via western blot. Autophagosome was investigated under a transmission electron microscope. After co-treatment with autophagy inhibitor Bafilomycin A1 or si-Beclin-1, cell apoptosis was detected by flow cytometry. Furthermore, under cotreatment with PI3K activator 740Y-P, PI3K/Akt/m-TOR pathway- and autophagy-related proteins were examined by western blot. Results: With a concentration-dependent manner, Propofol promoted the viability of HUVECs exposed to ox-LDL, and increased LC3-II/I ratio and Beclin-1 expression, and decreased P62 expression. The formation of autophagosome was enhanced by Propofol. Furthermore, Propofol treatment elevated Bcl-2/Bax ratio and lowered Caspase-3 expression. Bafilomycin A1 or si-Beclin-1 distinctly ameliorated the inhibitory effects of Propofol on apoptosis in ox-LDL-exposed HUVECs. Moreover, Propofol lowered the activation of PI3K/Akt/m-TOR pathway in HUVECs under exposure to ox-LDL. However, its inhibitory effects were weakened by 740Y-P. Conclusion: Collectively, this study revealed that Propofol could ameliorate ox-LDL-induced endothelial damage through enhancing autophagy via PI3K/Akt/m-TOR pathway, which might offer a novel therapeutic strategy in AS.

Propofol Mediated Protection of the Brain From Ischemia/Reperfusion Injury Through the Regulation of Microglial Connexin 43.

Cerebral ischemia/reperfusion (I/R) injury is a serious condition that leads to increased apoptosis of microglial and neurons in the brain. In this study, we identified that Cx43 expression level is significantly increased in the microglial cells during I/R injury. Using an in vitro model (hypoxia/reoxygenation-H/R injury), we observed that H/R injury leads to an increase in activation of microglial cells and increase in levels of pro-inflammatory markers such as IL-1β, IL-6, and TNF-α. Additionally, we could also observe significant increase in phosphorylation of Cx43 and Cav3.2 levels. To assess the role of H/R injured microglial cells on neuronal population, we cultured the neurons with conditioned media (MCS) from H/R injured microglial cells. Interestingly, we observed that microglial H/R injury significantly decreased Map2 expression and affected neuronal morphology. Further, we aimed to assess the effects of propofol on cerebral H/R injury, and observed that 40 μM propofol significantly decreased Cx43, Cx43 phosphorylation, and CaV3.2 levels. Additionally, propofol decreased apoptosis and increased Map2 expression levels in H/R injured neurons. Using silencing experiments, we confirmed that siCx43 could significantly improve the propofol’s rescue after H/R injury in both microglia and neurons. We further developed an in vivo MCAO (middle cerebral artery occlusion) rat model to understand the effect of propofol in I/R injury. Interestingly, propofol treatment and downregulation of Cx43 significantly decreased the infract volume and apoptosis in these MCAO rats. Thus, this study clearly establishes that propofol protects the brain against I/R injury through the downregulation of Cx43 in microglial cells.

Effects of propofol on the proliferation and migration of liver cancer cells.

Liver cancer is a malignant cancer with worldwide prevalence. It has been reported that cancer cells are usually exposed to a hypoxic microenvironment, which is associated with a poor prognosis in patients with cancer. Propofol is an intravenous anesthetic that is widely used in cancer surgery. The present study aimed to determine the effects of propofol stimulation on the viability, proliferation and migration of liver cancer cells under normoxia and cobalt chloride (CoCl2)-induced hypoxia. Under normoxia, HepG2 and HCCLM3 cells were randomly divided into six groups as follows: i) Control group; ii) 10 µM propofol group; iii) 25 µM propofol group; iv) 50 µM propofol group; v) 100 µM propofol group; and vi) DMSO group. Cell viability and proliferation were analyzed using Cell Counting Kit-8 (CCK-8) and 5-ethynyl-2'-deoxyuridine (EdU) assays, respectively, following 24 or 48 h of propofol treatment. In addition, wound healing and Transwell migration assays were used to determine the changes in cell migration. Under CoCl2-induced hypoxia, the protein levels of hypoxia inducible factor-1α (HIF-1α) of HepG2 cells were analyzed using western blotting. Subsequently, CCK-8 and wound healing assays were used to determine the effect of propofol on cell viability and migration. The results of the present study revealed that propofol stimulation had no significant effect on the viability, proliferation and migration of HepG2 and HCCLM3 cells under normoxia. The protein levels of HIF-1α were significantly upregulated following the treatment with 200 µM CoCl2 for 12 h. However, no significant differences were found in the viability and migration of HepG2 cells following the stimulation with propofol in the presence of CoCl2. In conclusion, the findings of the present study revealed that propofol exerted no effect on the viability, proliferation and migration of HepG2 and HCCLM3 cells under normoxic and hypoxic conditions.

This Month in Anesthesiology

This Month in Anesthesiology

Fast-spiking Interneurons Contribute to Propofol-induced Facilitation of Firing Synchrony in Pyramidal Neurons of the Rat Insular Cortex.

The general anesthetic propofol induces frontal alpha rhythm in the cerebral cortex at a dose sufficient to induce loss of consciousness. The authors hypothesized that propofol-induced facilitation of unitary inhibitory postsynaptic currents would result in firing synchrony among postsynaptic pyramidal neurons that receive inhibition from the same presynaptic inhibitory fast-spiking neurons. Multiple whole cell patch clamp recordings were performed from one fast-spiking neuron and two or three pyramidal neurons with at least two inhibitory connections in rat insular cortical slices. The authors examined how inhibitory inputs from a presynaptic fast-spiking neuron modulate the timing of spontaneous repetitive spike firing among pyramidal neurons before and during 10 μM propofol application. Responding to activation of a fast-spiking neuron with 150-ms intervals, pyramidal cell pairs that received common inhibitory inputs from the presynaptic fast-spiking neuron showed propofol-dependent decreases in average distance from the line of identity, which evaluates the coefficient of variation in spike timing among pyramidal neurons: average distance from the line of identity just after the first activation of fast-spiking neuron was 29.2 ± 24.1 (mean ± SD, absolute value) in control and 19.7 ± 19.2 during propofol application (P < 0.001). Propofol did not change average distance from the line of identity without activating fast-spiking neurons and in pyramidal neuron pairs without common inhibitory inputs from presynaptic fast-spiking neurons. The synchronization index, which reflects the degree of spike synchronization among pyramidal neurons, was increased by propofol from 1.4 ± 0.5 to 2.3 ± 1.5 (absolute value, P = 0.004) and from 1.5 ± 0.5 to 2.2 ± 1.0 (P = 0.030) when a presynaptic fast-spiking neuron was activated at 6.7 and 10 Hz, respectively, but not at 1, 4, and 13.3 Hz. These results suggest that propofol facilitates pyramidal neuron firing synchrony by enhancing inhibitory inputs from fast-spiking neurons. This synchrony of pyramidal neurons may contribute to the alpha rhythm associated with propofol-induced loss of consciousness.

Propofol activates AMPK to inhibit the growth of HepG2 cells in vitro and hepatocarcinogenesis in xenograft mouse tumor models by inducing autophagy

Hepatocellular carcinoma (HCC) is a fatal malignant tumor with a poor prognosis, and is the third leading cause of cancer-related deaths worldwide. This study aimed to investigate the anti-tumor effect of propofol on the proliferation, apoptosis, and cell cycle of HCC by regulating adenosine monophosphate-activated protein kinase (AMPK) in vivo and in vitro. The cell counting Kit-8 (CCK-8) assay was employed to screen the effect of propofol on HepG2 cell viability at various concentrations (0.3, 0.6, 1.2, 2.5, 5, 10, 20, 40, 80 and 160 µM). We selected propofol at concentrations of 5, 10 and 20 µM for subsequent experiments. Flow cytometry was used to examine the apoptosis and cell cycle of HCC. Quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) was applied to measure the messenger ribonucleic acid (mRNA) expression levels of proliferating cell nuclear antigen (PCNA) and survivin. Western blotting was applied to measure the protein expression levels of PCNA, survivin, cleaved caspase-3, cleaved caspase-9, p27 (Kip1), and cyclin A. The effects of propofol were evaluated by establishing a xenograft tumor model. After treatment with propofol, the mRNA expression levels of PCNA and survivin were decreased compared with the 0 µM propofol (control) group. The colony formation assay showed that the colony formation rate was obviously down-regulated. Flow cytometry demonstrated that HepG2 cell apoptosis was increased. G0/G1 was enhanced compared with the control group, while G2/M was restrained. The levels of cleaved caspase-3, cleaved caspase-9, p27, phospho-AMP-activated protein kinase α1 (p-AMPKα1), phospho-mammalian target of rapamycin (p-mTOR), and phospho-Unc-51 like autophagy activating kinase 1 (p-ULK1) were notably elevated, while the levels of cyclin A were suppressed. The xenograft tumor volume declined in vivo compared with the HepG2 xenograft group. The expression levels of cell proliferation markers (PCNA) were significantly down-regulated markedly, while the expression levels of cell cycle markers (p27) were notablyup-regulated. Terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) staining showed that cell apoptosis was increased. The levels of p-AMPKα1 were also up-regulated. Propofol inhibits the proliferation, apoptosis, and cell cycle of HCC by regulating AMPK in vivo and in vitro.

Membrane nanodomains homeostasis during propofol anesthesia as function of dosage and temperature

Some anesthetics bind and potentiate γ-aminobutyric-acid-type receptors, but no universal mechanism for general anesthesia is known. Furthermore, often encountered complications such as anesthesia induced amnesia are not understood. General anesthetics are hydrophobic molecules easily dissolving into lipid bilayers. Recently, it was shown that general anesthetics perturb phase separation in vesicles extracted from fixed cells. Unclear is whether under physiological conditions general anesthetics induce perturbation of the lipid bilayer, and whether this contributes to the transient loss of consciousness or anesthesia side effects. Here we show that propofol perturbs lipid nanodomains in the outer and inner leaflet of the plasma membrane in intact cells, affecting membrane nanodomains in a concentration dependent manner: 1 μM to 5 μM propofol destabilize nanodomains; however, propofol concentrations higher than 5 μM stabilize nanodomains with time. Stabilization occurs only at physiological temperature and in intact cells. This process requires ARP2/3 mediated actin nucleation and Myosin II activity. The rate of nanodomain stabilization is potentiated by GABAA receptor activity. Our results show that active nanodomain homeostasis counteracts the initial disruption causing large changes in cortical actin. Significance statementGeneral anesthesia is a routine medical procedure with few complications, yet a small number of patients experience side-effects that persist for weeks and months. Very young children are at risk for effects on brain development. Elderly patients often exhibit subsequent amnesia. Here, we show that the general anesthetic propofol perturbs the ultrastructure of the lipid bilayer of the cell membrane in intact cells. Initially propofol destabilized lipid nanodomains. However, with increasing incubation time and propofol concentration, the effect is reversed and nanodomains are further stabilized. We show that this stabilization is caused by the activation of the actin cortex under the membrane. These perturbations of membrane bilayer and cortical actin may explain how propofol affects neuronal plasticity at synapses.

MiR-455-3p alleviates propofol-induced neurotoxicity by reducing EphA4 expression in developing neurons

Purpose Propofol, an aesthetic agent in paediatric patients, results in neurotoxicity in the developing neurons. To reduce side effects of propofol, the protective role of miR-455-3p (microRNA-455-3p) in developing rat brain was investigated. Materials and methods Primary hippocampal neurons were isolated from postnatal day 1 or 2 SD (Sprague-Dawley) rats. The neurons were exposed to various concentrations of propofol (0, 10, 30, or 50 μM) for 6 h. Propofol-induced cell viability was assessed by MTT assay, expression levels of miR-455-3p and EphA4 (erythropoietin-producing hepatocellular A4) in propofol-induced neurons were determined using qRT-PCR and western blot, respectively. Binding ability between miR-455-3p and EphA4 was predicted, and then validated by luciferase reporter assay. Neurons expressing miR-455-3p mimics, were treated with 50 μM propofol for 6 h, and apoptosis status was evaluated by flow cytometry. Results Exposure to propofol significantly decreased cell viability of developing neurons isolated from neonatal rats. Propofol decreased miR-455-3p expression, while increased EphA4 level in the neurons. miR-455-3p mimics increased propofol-induced reduce in cell viability, and attenuated propofol-induced cell apoptosis of neurons. MiR-455-3p could target EphA4, and decreased expression of EphA4 in neurons exposure to propofol. EphA4 knockdown counteracted with the promotive effects of propofol on cell viability and apoptosis of neurons. Conclusion Propofol treatment induces neurotoxicity and suppresses miR-455-3p levels in the developing hippocampal neurons. However, miR-455-3p could alleviate such neurotoxicity by reducing EphA4 expression, provided new insights into miR-455-3p as novel therapeutic target to prevent propofol-induced damages from bench to clinic.

Allopregnanolone Enhances GABAergic Inhibition in Spinal Motor Networks.

The neurosteroid allopregnanolone (ALLO) causes unconsciousness by allosteric modulation of γ-aminobutyric acid type A (GABAA) receptors, but its actions on the spinal motor networks are unknown. We are therefore testing the hypothesis that ALLO attenuates the action potential firing of spinal interneurons and motoneurons predominantly via enhancing tonic, but not synaptic GABAergic inhibition. We used video microscopy to assess motoneuron-evoked muscle activity in organotypic slice cultures prepared from the spinal cord and muscle tissue. Furthermore, we monitored GABAA receptor-mediated currents by performing whole-cell voltage-clamp recordings. We found that ALLO (100 nM) reduced the action potential firing of spinal interneurons by 27% and that of α-motoneurons by 33%. The inhibitory effects of the combination of propofol (1 µM) and ALLO on motoneuron-induced muscle contractions were additive. Moreover, ALLO evoked a tonic, GABAA receptor-mediated current (amplitude: 41 pA), without increasing phasic GABAergic transmission. Since we previously showed that at a clinically relevant concentration of 1 µM propofol enhanced phasic, but not tonic GABAergic inhibition, we conclude that ALLO and propofol target distinct subpopulations of GABAA receptors. These findings provide first evidence that the combined application of ALLO and propofol may help to reduce intraoperative movements and undesired side effects that are frequently observed under total intravenous anesthesia.

Effects of Propofol on Electrical Synaptic Strength in Coupling Reticular Thalamic GABAergic Parvalbumin-Expressing Neurons.

Electrical synapses between neurons exhibit a high degree of plasticity, which makes critical contributions to neuronal communication. The GABAergic parvalbumin-expressing (PV+) neurons in the thalamic reticular nucleus (TRN) interact with each other through electrical and chemical synapses. Plasticity of electrical synaptic transmission in TRN plays a key role in regulating thalamocortical and corticothalamic circuits and even the formation of consciousness. We here examined the effects of propofol, a commonly used general anesthetic agent, on the strength of electrical synapses between TRN PV+ neurons by fluorescence-guided patch-clamp recording and pharmacological methods. Results show that 100 μM propofol reduced the electrical synaptic strength between TRN PV+ neurons. Notably, the propofol-induced depression of electrical synaptic strength between TRN PV+ neurons was diminished by saclofen (10 μM, antagonist of GABAB receptors), but not blocked by gabazine (10 μM, antagonist of GABAA receptors). Application of baclofen (10 μM, agonist of GABAB receptors), similar to propofol, also reduced the electrical synaptic strength between TRN PV+ neurons. Moreover, the propofol-induced depression of electrical synaptic strength between TRN PV+ neurons was abolished by 9-CPA (100 μM, specific adenylyl cyclase inhibitor), and by KT5720 (1 μM, selective inhibitor of PKA). Our findings indicate that propofol acts on metabotropic GABAB receptors, resulting in a depression of electrical synaptic transmission of coupled TRN PV+ neurons, which is mediated by the adenylyl cyclase-cAMP-PKA signaling pathway. Our findings also imply that propofol may change the thalamocortical communication via inducing depression of electrical synaptic strength in the TRN.

Propofol Reduces Epithelial to Mesenchymal Transition, Invasion and Migration of Gastric Cancer Cells through the MicroRNA-195-5p/Snail Axis.

BackgroundGastric cancer (GC) is a life-threating malignancy worldwide. Accumulating studies suggest propofol has anti-tumor functions in addition to the anesthetic effect. This study aimed to figure out the effects of propofol treatment in GC development.Material/MethodsHuman GC SGC-7901 and NCI-N87 cells were treated with different doses of propofol. Then the invasion and migration of GC cells was measured. SGC-7901 cells following 10 μM propofol treatment were applied in the following experiments. MicroRNAs (miRNAs) with differential expression in cells with or without propofol treatment were analyzed. Expression of miR-195-5p, Snail, vimentin and E-cadherin in SGC-7901 cells was measured, and then loss-of-function of miR-195-5p and gain-of-function of Snail were performed. Target relation between miR-195-5p and Snail was confirmed using luciferase assay. Xenograft tumor was induced in nude mice to identify the effect of propofol on GC in vivo.ResultsPropofol reduced epithelial to mesenchymal transition (EMT), invasion and migration of GC cells in a dose-dependent manner. Propofol elevated miR-195-5p expression but reduced Snail expression, and it reduced vimentin but increased E-cadherin expression in SGC-7901 cells. miR-195-5p directly bound to Snail. miR-195-5p inhibition or Snail promotion reversed propofol-inhibited malignant behaviors of SGC-7901 cells. In vitro results were reproduced in in vivo experiments.ConclusionsOur study found that propofol could inhibit EMT, invasion, and migration of GC cells by promoting miR-195-5p expression and suppressing Snail expression. This study may provide novel insights in GC treatment.

Propofol improves colonic but impairs hepatic mitochondrial function in tissue homogenates from healthy rats

Evidence suggests that propofol infusion syndrome (PRIS) is caused by an altered mitochondrial function. The aim of this study was to examine the effects of propofol and the vehicle MCT on mitochondrial function in hepatic and colonic tissue.Mitochondrial oxygen consumption was determined in colon and liver homogenates after incubation with buffer (control), propofol (50, 75, 100, 500 μM) or the carrier substances DMSO and MCT. State 2 (substrate-dependent) and state 3 (ADP-dependent respiration) were assessed. RCI (respiratory control index) - an indicator for coupling between electron transport chain system (ETS) and oxidative phosphorylation (OXPHOS) and ADP/O ratio - a parameter for efficacy of OXPHOS were calculated. Data were presented as % of control.In hepatic mitochondria, 500 μM propofol reduced RCI formulation-independently (propofol/MCT 500 μM: complex I: 66.3 ± 8.7%*, complex II: 75.5 ± 9.2%*; propofol/DMSO 500 μM: complex I: 29.1 ± 8.8%*, complex II: 49.3 ± 15.5%*). 75 μM Propofol/MCT reduced ADP/O for complex I (73.5 ± 27.3%*). DMSO did not affect hepatic mitochondria whereas MCT reduced RCI for complex II (87.2 ± 9.8%*) and ADP/O for complex I (93.7 ± 31.7%*). In colon 50 μM Propofol/MCT increased RCI for complex I and II (complex I: 127.2 ± 10.7%*, complex II: 136.8 ± 33.9%*) and 100 μM Propofol/MCT for complex I (131.4 ± 18.7%*). 500 μM Propofol/DMSO increased ADP/O for complex I (139.4 ± 41.4%*). DMSO did not affect RCI but increased ADP/O for both complexes (complex I: 119.9 ± 25.8%*, complex II: 110.2 ± 14.2%*). MCT increased RCI for complex I (123.0 ± 31.6%*).In hepatic mitochondria propofol uncoupled ETS from OXPHOS formulation-independently and propofol/MCT reduced efficacy of OXPHOS. In colonic mitochondria, propofol/MCT strengthened the coupling and propofol/DMSO enhanced the efficacy of OXPHOS.