Membrane Lipids and Osmolytes in the Response of the Acidophilic Basidiomycete Phlebiopsis gigantea to Heat, Cold, and Osmotic Shocks.

Previously, we found for the first time the participation of osmolytes in adaptation to acidic conditions in three acidophilic fungi. Because trehalose can protect membranes, we hypothesized a relationship between osmolyte and membrane systems in adaptation to stressors. In the mycelium of Phlebiopsis gigantea, the level of osmolytes reaches 8% of the dry mass, while trehalose and arabitol make up 60% and 33% of the sum, respectively. Cold shock does not change the composition of osmolytes, heat shock causes a twofold increase in the trehalose level, and osmotic shock leads to a marked increase in the amount of trehalose and arabitol. Predominance of phospholipids (89% of the sum) and low proportions of sterols and sphingolipids are characteristic features of the membrane lipids' composition. Phosphatidic acids, along with phosphatidylethanolamines and phosphatidylcholines, are the main membrane lipids. The composition of the membrane lipids remains constant under all shocks. The predominance of linoleic (75% of the sum) and palmitic (20%) acids in phospholipids results in a high degree of unsaturation (1.5). Minor fluctuations in the fatty acid composition are observed under all shocks. The results demonstrate that maintaining or increasing the trehalose level provides stability in the membrane lipid composition during adaptation.

Tissue Lipidomic Alterations Induced by Prolonged Dexamethasone Treatment.

Dexamethasone is a synthetic glucocorticoid medication vastly used to treat abnormal immune responses and inflammation. Although the medication is well-established in the medical community, the prolonged treatment with high dosages of dexamethasone may lead to severe adverse effects through mechanisms that are not yet well-known. Lipids are a large class of hydrophobic molecules involved in energy storage, signaling, modulation of gene expression, and membranes. Hence, untargeted lipidomics may help unravel the biochemical alterations following prolonged treatment with high dosages of dexamethasone. We performed comprehensive lipidomic analyses of brain, heart, kidney, liver, and muscle samples obtained from rats that were treated with intramuscular injections of dexamethasone for 14 weeks compared to healthy controls. The employed methodology and statistical analysis showed that phosphatidic acids, glycerophospholipids, plasmalogens, and fatty acids are deeply affected by prolonged use of the medication. Brain tissue was only mildly affected, but skeletal muscle showed a strong accumulation of lipids that may be correlated with alterations in the energy metabolism, myopathy, and oxidative processes. This work provides new insights into the mechanisms of action and adverse effects for one of the most commonly prescribed class of drugs in the world.

Effect of humic acid on the composition of osmolytes and lipids in a melanin-containing phytopathogenic fungus Alternaria alternata.

Humic substances (HS) have a direct impact on living cells, causing a wide range of various biological effects, and stimulating or inhibiting fungal growth. Therefore, it is important to reveal cellular indicators that could indicate the nature and level of HS' effects on living organisms. Paying attention to the important role of lipid and osmolytes in adaptation to stress, the aim of this work was to study the composition of osmolytes and lipids in the growth dynamics of the phytopathogenic melanin-containing fungus Alternaria alternata under the influence of humic acid (HA). HA was isolated from a commercial peat humic product. For cultivation, liquid Czapek medium with the addition of 200 mg/l HA (HA variant) was used, and with no HA for the control variant. The main osmolytes of the fungal mycelium were glucose, mannitol, and trehalose. Both in the control and in HA variants, a general pattern was observed during the growth process; the amount of glucose increased against the background of a decrease of mannitol. In the control, the amount of trehalose increased significantly by the stage of active melanin formation (day 4), while under the influence of HA, such an increase was not observed. Membrane lipids were represented by phospholipids, sphingolipids, and sterols. A characteristic feature of A. alternata was a high proportion of phosphatidic acids (PA) in the composition of membrane lipids. The amount and proportion of PA decreased during growth in the control, while under the influence of HA such decline was not observed. The influence of HA induced the increase in the amount of sterols (St) and phosphatidylcholines (PC) that were able to stabilise the membrane lipid bilayer. We can assume that general cellular response of A. alternata to HA influence, being expressed as a higher amount of PA, PC, St and a lower level of trehalose in comparison with the control, indicates the an absence of inhibition of growth processes of this phytopathogenic melanin-producing fungus. The effect of HA on the qualitative and quantitative composition of the osmolytes and membrane lipids of A. alternata may cause changes of virulence and stress resistance of this phytopathogen.

Combinatorial impact of osmotic and heat shocks on the composition of membrane lipids and osmolytes in Aspergillus niger.

The combinatorial action of osmotic (OS) and heat (HS) shocks on the composition of soluble cytosol carbohydrates and membrane lipids was studied. For the first time it was demonstrated that the combinatorial effect of these shocks led to the non-additive response - an increase in the trehalose level, characteristic for HS, but at the same time suppression of glycerol production, uncharacteristic of the OS response. In addition, combinatorial action resulted in a new effect - increase in the mannitol level, which was not typical for the individual HS or OS responses. On the contrary, a general pattern of change was observed in the composition of membrane lipids in response to both individual HS and OS, and their combinations, which was a twofold increase in the proportion of phosphatidic acids. At the same time, the mechanism of alteration in the degree of unsaturation of membrane phospholipids was not involved in adaptation. The response to combinatorial shocks includes the accumulation of trehalose and mannitol, and increase in the proportion of phosphatidic acids in membrane lipids.

Topological organisation of the phosphatidylinositol 4,5-bisphosphate-phospholipase C resynthesis cycle: PITPs bridge the ER-PM gap.

Phospholipase C (PLC) is a receptor-regulated enzyme that hydrolyses phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) at the plasma membrane (PM) triggering three biochemical consequences, the generation of soluble inositol 1,4,5-trisphosphate (IP3), membrane-associated diacylglycerol (DG) and the consumption of PM PI(4,5)P2 Each of these three signals triggers multiple molecular processes impacting key cellular properties. The activation of PLC also triggers a sequence of biochemical reactions, collectively referred to as the PI(4,5)P2 cycle that culminates in the resynthesis of this lipid. The biochemical intermediates of this cycle and the enzymes that mediate these reactions are topologically distributed across two membrane compartments, the PM and the endoplasmic reticulum (ER). At the PM, the DG formed during PLC activation is rapidly converted into phosphatidic acid (PA) that needs to be transported to the ER where the machinery for its conversion into PI is localised. Conversely, PI from the ER needs to be rapidly transferred to the PM where it can be phosphorylated by lipid kinases to regenerate PI(4,5)P2 Thus, two lipid transport steps between membrane compartments through the cytosol are required for the replenishment of PI(4,5)P2 at the PM. Here, we review the topological constraints in the PI(4,5)P2 cycle and current understanding how these constraints are overcome during PLC signalling. In particular, we discuss the role of lipid transfer proteins in this process. Recent findings on the biochemical properties of a membrane-associated lipid transfer protein of the PITP family, PITPNM proteins (alternative name RdgBα/Nir proteins) that localise to membrane contact sites are discussed. Studies in both Drosophila and mammalian cells converge to provide a resolution to the conundrum of reciprocal transfer of PA and PI during PLC signalling.

Cancer cells incorporate and remodel exogenous palmitate into structural and oncogenic signaling lipids.

De novo lipogenesis is considered the primary source of fatty acids for lipid synthesis in cancer cells, even in the presence of exogenous fatty acids. Here, we have used an isotopic fatty acid labeling strategy coupled with metabolomic profiling platforms to comprehensively map palmitic acid incorporation into complex lipids in cancer cells. We show that cancer cells and tumors robustly incorporate and remodel exogenous palmitate into structural and oncogenic glycerophospholipids, sphingolipids, and ether lipids. We also find that fatty acid incorporation into oxidative pathways is reduced in aggressive human cancer cells, and instead shunted into pathways for generating structural and signaling lipids. Our results demonstrate that cancer cells do not solely rely on de novo lipogenesis, but also utilize exogenous fatty acids for generating lipids required for proliferation and protumorigenic lipid signaling. This article is part of a special issue entitled Lipid Metabolism in Cancer.

Acquired thermotolerance, membrane lipids and osmolytes profiles of xerohalophilic fungus Aspergillus penicillioides under heat shock.

Xerophilic fungi accumulate a large amount of glycerol in the cytosol to counterbalance the external osmotic pressure. But during heat shock (HS) majority of fungi accumulate a thermoprotective osmolyte trehalose. Since glycerol and trehalose are synthesized in the cell from the same precursor (glucose), we hypothesised that, under heat shock conditions, xerophiles growing in media with high concentrations of glycerol may acquire greater thermotolerance than those grown in media with high concentrations of NaCl. Therefore, the composition of membrane lipids and osmolytes of the fungus Aspergillus penicillioides, growing in 2 different media under HS conditions was studied and the acquired thermotolerance was assessed. It was found that in the salt-containing medium an increase in the proportion of phosphatidic acids against a decrease in the proportion of phosphatidylethanolamines is observed in the composition of membrane lipids, and the level of glycerol in the cytosol decreases 6-fold, while in the medium with glycerol, changes in the composition of membrane lipids are insignificant and the level of glycerol is reduced by no more than 30%. In the mycelium trehalose level have increased in both media, but did not exceed 1% of dry weight. However, after exposure to HS the fungus acquires greater thermotolerance in the medium with glycerol than in the medium with salt. The data obtained indicate the interrelation between changes in the composition of osmolytes and membrane lipids in the adaptive response to HS, as well as the synergistic effect of glycerol and trehalose.

The Role of Osmolytes and Membrane Lipids in the Adaptation of Acidophilic Fungi.

Acidophiles maintain near-neutral intracellular pH using proton pumps. We have suggested the protective role of osmolytes and membrane lipids in the adaptation to an acidic environment. Previously we have observed, for the first time, high levels of trehalose in acidophilic basidiomycete Sistotrema brinkmannii. Here, we have studied the composition of both osmolytes and membrane lipids of two more acidophilic fungi. Trehalose and polyols were among the main osmolytes during growth under optimal conditions (pH 4.0) in basidiomycete Phlebiopsis gigantea and ascomycete Mollisia sp. Phosphatidic acids, phosphatidylethanolamines, phosphatidylcholines, and sterols, were predominant membrane lipids in both fungi. P. gigantea had a narrow optimum of growth at pH 4.0, resulting in a sharp decline of growth rate at pH 2.6 and 5.0, accompanied by a decrease in the number of osmolytes and significant changes in the composition of membrane lipids. In contrast, Mollisia sp. had a broad optimal growth range (pH 3.0-5.0), and the number of osmolytes either stayed the same (at pH 6.0) or increased (at pH 2.6), while membrane lipids composition remained unchanged. Thus, the data obtained indicate the participation of osmolytes and membrane lipids in the adaptation of acidophilic fungi.

Natural exosome-like nanovesicles from edible tea flowers suppress metastatic breast cancer via ROS generation and microbiota modulation.

Although several artificial nanotherapeutics have been approved for practical treatment of metastatic breast cancer, their inefficient therapeutic outcomes, serious adverse effects, and high cost of mass production remain crucial challenges. Herein, we developed an alternative strategy to specifically trigger apoptosis of breast tumors and inhibit their lung metastasis by using natural nanovehicles from tea flowers (TFENs). These nanovehicles had desirable particle sizes (131 nm), exosome-like morphology, and negative zeta potentials. Furthermore, TFENs were found to contain large amounts of polyphenols, flavonoids, functional proteins, and lipids. Cell experiments revealed that TFENs showed strong cytotoxicities against cancer cells due to the stimulation of reactive oxygen species (ROS) amplification. The increased intracellular ROS amounts could not only trigger mitochondrial damage, but also arrest cell cycle, resulting in the in vitro anti-proliferation, anti-migration, and anti-invasion activities against breast cancer cells. Further mice investigations demonstrated that TFENs after intravenous (i.v.) injection or oral administration could accumulate in breast tumors and lung metastatic sites, inhibit the growth and metastasis of breast cancer, and modulate gut microbiota. This study brings new insights to the green production of natural exosome-like nanoplatform for the inhibition of breast cancer and its lung metastasis via i.v. and oral routes.

Arabidopsis leaf extracellular vesicles in wound-induced jasmonate accumulation.

The plant extracellular vesicles (EVs) are lipid-enveloped nano-particles containing proteins, nucleic acids and metabolites and function in plant development and response. The Arabidopsis four transmembrane protein TETRASPANIN 8 (TET8) knock-out mutant tet8 secreted less EVs than the wild-type (WT). In this report, we show that the tet8 mutant was attenuated in the plant hormone jasmonate (JA) accumulation in response to mechanical wounding treatment. We also noticed that the EVs contained a high level of phospholipids phosphatidic acids (PAs) which may serve as precursors of JA biosynthesis during wound-triggered-self-healing processes. Thus, we propose an open question about a potential role of EVs or TET8 or both in damage-associated JA response.

Osmolytes and membrane lipids in the adaptation of micromycete Emericellopsis alkalina to ambient pH and sodium chloride.

The accumulation of low molecular weight cytoprotective compounds (osmolytes) and changes in the membrane lipids composition are of key importance for the adaptation to stress impacts. However, the reason behind the wide variety of osmolytes present in the cell remains unclear. We suggest that specific functions of osmolytes can be revealed by studying the adaptation mechanisms of the mycelial fungus Emericellopsis alkalina (Hypocreales, Ascomycota) that is resistant to both alkaline pH values and high sodium chloride concentrations. It has been established that the fungus uses different osmolytes to adapt to ambient pH and NaCl concentration. Arabitol was predominant osmolyte in alkaline conditions, while mannitol prevailed in acidic conditions. On the salt-free medium mannitol was the main osmolyte; under optimal conditions (pH 10.2; 0.4 M NaCl) arabitol and mannitol were both predominant. Higher NaCl concentrations (1.0-1.5 M) resulted in the accumulation of low molecular weight polyol - erythritol, which amounted up to 12-14%, w/w. On the contrary, changes in the composition of membrane lipids were limited under pH and NaCl impacts; only higher NaCl concentrations led to the increase in the degree of unsaturation of membrane lipids. Results obtained indicated the key role of the osmolytes in the adaptation to the ambient pH and osmotic impacts.

Phospholipases D: making sense of redundancy and duplication.

Why have two genes when one would suffice? Evolutionary pressure means that biology, unlike government, is generally intolerant of wasted effort. Therefore, when multiple genes exist presumably they are there to provide some benefit to the organism even if that benefit is not immediately obvious to us scientists. A recent report from Raghu and colleagues (Biosci. Rep. (2018) 38, pii: BSR20181690) [1] sheds some light on one possible reason for the existence of two Phospholipases D genes in chordates when only one is present in invertebrates.

Lipids in Regulated Exocytosis: What are They Doing?

The regulated secretory pathway in neuroendocrine cells ends with the release of hormones and neurotransmitters following a rise in cytosolic calcium. This process known as regulated exocytosis involves the assembly of soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins, the synaptic vesicle VAMP (synaptobrevin), and the plasma membrane proteins syntaxin and SNAP-25. Although there is much evidence suggesting that SNARE proteins play a key role in the fusion machinery, other cellular elements regulating the kinetics, the extent of fusion, and the preparation of vesicle for release have received less attention. Among those factors, lipids have also been proposed to play important functions both at the level of secretory vesicle recruitment and late membrane fusion steps. Here, we will review the latest evidence supporting the concept of the fusogenic activity of lipids, and also discuss how this may be achieved. These possibilities include the recruitment and sequestration of the components of the exocytotic machinery, regulation of protein function, and direct effects on membrane topology.

Clinical effect of alprostadil combined with Kudiezi injection in the treatment of posterior circulation ischemic vertigo and its effect on levels of Iysophosphatidic acid, acidic phospholipid / 中国基层医药

Objective To explore the clinical effect of alprostadil combined with Kudiezi injection in the treatment of posterior circulation ischemic vertigo,and its effect on levels of lysophosphatidic acid (LPA),acidic phospholipid (AP).Methods From October 2015 to October 2017,92 cases of posterior circulation ischemia in the Affiliated Hospital of Medical College were selected and randomly divided into observation group(n =46) and control group(n =46) according to the digital table.The control group was treated with Kudiezi injection,while the observation group was treated with alprostadil combined with Kudianzi injection.The clinical efficacy and LPA,AP levels before and after treatment were compared between the two groups.Results After treatment,the total effective rate in the observation group was 95.65％,which in the control group was 82.61％,there was statistically significant difference between the two groups(x2 =8.622,P ＜0.05).After treatment,Vm and Vs of bilateral vertebrobasilar artery in both two groups were increased more rapidly than those before treatment(observation group:t =14.041,11.124,11.207,10.057,10.925,11.920;control group:t =7.204,7.057,8.145,6.572,6.581,5.481,all P ＜ 0.05).Compared with the control group,the Vm [(34.24 ± 3.04) cm/s,(30.54 ± 3.33) cm/s,(35.42 ± 3.46) cm/s] and Vs[(40.09 ± 5.14) cm/s,(40.24 ± 5.02) cm/s,(43.14 ± 4.97) cm/s] of bilateral vertebrobasilar artery in the observation group were significantly higher (t =7.825,4.581,8.610,7.256,7.017,5.824,all P ＜ 0.05).After treatment,the levels of LPA and AP in the two groups were significantly lower than those before treatment(observation group:t =18.054,17.259;control group:t =17.651,14.254,all P ＜ 0.05).The levels of LPA and AP in the control group [(1.75 ± 0.52) μmol/L,(2.42 ± 0.51) μmol/L] were significantly higher than those in the observation group [[(1.05 ± 0.28) μmol/L,(1.84 ± 0.48) μmol/L] (t =8.571,7.224,all P ＜ 0.05).Before treatment,the number of white blood cells in two groups were (6.23 ±0.54) × 109/L,(6.68 ±0.57) × 109/L,respectively,which after treatment were (6.57 ±0.61) × 109/L,(6.42 ±0.64) × 109/L,respectively,there was no statistically significant difference in leukocyte count between the two groups before and after treatment(all P ＜ 0.05).During the treatment,there was no obvious adverse reaction in the two groups.Conclusion Alprostadil combined with Kudiezi injection in the treatment of circulatory ischemic vertigo has excellent clinical effect,there are no adverse reactions such as leukopenia occurred and the safety is good.

Clinical study on treatment of Bushen-Huoxue decoction for the patients with senile chronic cerebral circulation insufficiency / 国际中医中药杂志

Objective To observe the effect of Bushen-Huoxue decoction combined with conventional therapy on the clinical symptoms, vertigo degree and cognitive function of elderly patients with chronic cerebral cerebral circulation insufficiency (CCCI) and to explore its mechanism. Methods A total of 70 patients were randomly divided into observation group and control group according to random number table method. The two groups were given antihypertensive, hypoglycemic, lipid and other western medicine treatment. On this basic treatment, the control group added orally flunarizine hydrochloride capsules, 5 mg /day before sleeping, while the observation group with Bushen-Huoxue decoction per day. All the treatment last 30 days. dizziness handicap inventory (DHI) and montreal cognitive assessment (MoCA) were used for the symptoms evaluation, and the mean flow velocity of the following arteries basilar artery (BA), bilateral vertebral artery (VA) and middle cerebral artery (MCA) were assessed by the Ultrasound transcranial doppler blood flow analyzer. The acidity phosphatidic acid, AP were detected by chemical colorimetric method. And the clinical effect rates were compared after treatment. Results The total effective rate of observation group was 94.3％ (33/35), higher than 82.9％ (29/35) in the control group, and the difference in group 2 was statistically significant (χ2=9.728, P<0.05). After treatment, the TCM symptom scores (6.2 ± 3.3 vs. 9.8 ± 3.7, t=8.920), DHI score (4.4 ± 2.5 vs. 9.3 ± 3.6, t=12.081) and MoCA score (25.7 ± 2.6 vs. 23.2 ± 2.8, t=6.638) improvement of the observation group were significantly better than those of control group (P<0.05). The Vms of LVA (37.2 ± 8.5 cm/s vs. 34.9 ± 7.6 cm/s, t=9.103), LMCA (63.3 ± 9.8 cm/s vs. 60.1 ± 8.4 cm/s, t=7.839), RMCA (62.8 ± 10.5 cm/s vs. 60.9 ± 9.5 cm/s, t=6.583) and plasma AP (3.74 ± 1.08 μmol/L vs. 5.81 ± 1.35 μmol/L, t=9.627) improvement of the observation group were significantly better than those of control group (P<0.05). Conclusions The Bushen-Huoxue decoction can improve the clinical symptoms and vertigo of the elderly and improve the cognitive level of the patients. The mechanism may be related to the improvement of the cerebral blood flow velocity, the decrease of plasma AP, and the state of ischemia and hypoxia.

STAT3 is involved in phosphatidic acid-induced Bcl-2 expression in HeLa cells

Phosphatidic acid (PA), the product of a PLD-mediated reaction, is a lipid second messenger that participates in various intracellular signaling events and is known to regulate a growing list of signaling proteins. We found that Bcl-2 was upregulated by PA treatment in HeLa cells. However, how PA upregulates Bcl-2 expression has not yet been studied. In this study, we tried to discover the mechanisms of Bcl-2 up-regulation by PA treatment in HeLa cells. Treatment with PA resulted in significantly increased expression of Bcl-2 in HeLa cells. Moreover, PA-induced Bcl-2 expression was blocked by mepacrine, an inhibitor of PLA2, but not by propranolol, an inhibitor of PA phospholyhydrolase (PAP). Treatment of 1,2-dipalmitoryl-sn-glycero-3-phosphate (DPPA) also increased Bcl-2 expression. These results indicate that Bcl-2 expression is mediated by lysophosphatidic acid (LPA), not by arachidonic acid (AA). Thereafter, we used MEK1/2 inhibitor, PD98059 to investigate the relationship between ERK1/2 MAPK and PA-induced Bcl-2 expression. PA-induced Bcl-2 expression was decreased when ERK1/2 was inhibited by PD98059. The transcription factor such as STAT3 which is controlled by ERK1/2 MAPK was increased along with Bcl-2 expression when the cells were treated with PA. Furthermore, STAT3 siRNA treatments inhibited PA-induced Bcl-2 expression, suggesting that STAT3 (Ser727) is involved in PA-induced Bcl-2 expression. Taken together, these findings indicate that PA acts as an important mediator for increasing Bcl-2 expression through STAT3 (Ser727) activation via the ERK1/2 MAPK pathway.

Phosphatidic acid mediates inflammatory responses of macrophages derived from experimental glomerulonephritis / 中国病理生理杂志

AIM: To investigates the role of phosphatidic acid (PA) in the expression of several inflammatory mediators produced by glomerural macrophages (GM?). METHODS: The study was performed on a rat model of accelerated anti-glomerural basement membrane (anti-GBM) glomerulonephritis (GN). GN-GM? were isolated and identified. Peritoneal M? (P-M?) of both normal and GN rats were used as controls. Block and reverse test were investigated with rhIL-1? stimulated, lisofylline (LSF) and phosphatidic acid (PA). Macrophage expression of ICAM-1 and TGF-? 1 were assessed at the level of protein and gene by immunocytochemistry, northern blot and RT-PCR. RESULTS: (1) After stimulated with rhIL-1?, GN-GM? produced much more ICAM-1, MCP-1 and TGF-? 1 than P-M?, and it's gene expression was similar as protein product. (2) mRNA expression of these factors was up-regulated again after the GN-GM? were pretreated with LSF then PA was added. CONCLUSIONS: Since GN-GM? plays an important role for PA in the mediation of glomerular injury, inhibiting of PA production is the keypoint of blocking M? mediated inflammatory effects. LSF may be an effective medicine in therapy for acute inflammatory forms of GN.

The safety of inhalation of volatile anesthetic mixed with nitric oxide during mechanical ventilation in piglets / 中华麻醉学杂志

Objective To investigate the safety of inhalation of isoflurane (ISO) or sevoflurane (SEVO) mixed with nitric oxide (NO) during mechanical ventilation.Methods Thirty-six healthy piglets of both sexes weighing 7-11 kg were randomly allocated to one of 6 groups ( n = 6 each): (1) control group was mechanically ventilated with O2; (2) NO group inhaled 20 ppm NO; (3) ISO group inhaled 1.3 MAC isoflurane; (4) ISO + NO group 1.3 MAC isoflurane + 20 ppm NO; (5) SEVO group inhaled 1.3 MAC sevoflurane and (6) SEVO + NO group inhaled 1.3 MAC sevoflurane + 20 ppm NO. The animals were mechanically ventilated with IPPV (VT 10 ml?kg-1 , RR 30-40 bpm, I: E 1:2) for 4 h in the all 6 groups. The animals were premedicated with atropine 0.02 mg?kg-1 i.m. . The Ⅳ line was established for fluid and drug administration. An additional dose of ketamine 10 mg?kg-1 was given i.v. before tracheostomy. 4F S-G catheter was placed in pulmonary artery via right internal jugular vein for hemodynamic monitoring. Femoral artery was cannulated for BP monitoring and collection of artrerial blood samples. MAP, HR, CVP, right ventricular pressure (RVP), PCWP, MPAP and total compliance of respiratory system (Crs), Paw, VT and PET CO2 were recorded before (T0 ) and at 1, 2, 3, 4 h of ventilation (T1-4). Blood samples were taken at T0 , T2 and T4 for determination of Met Hb and NO2- /NO3- . The animals were killed at the end of 4 h mechanical ventilation and the lungs were removed for determination of wet/dry(W/D) lung weight ratio, broncho-alveolar lavage fluid (BALF) and microscopic examination. BALF was collected for determination of surface tension and concentrations of saturated phosphate (DSPC) , total phosphate (TPL) total protein (TP) and white cell count. Results Crs was significantly decreased at the end of ventilation (T4 ) as compared with the baseline (T0) in group 3,4,5 and 6, while there was no significant change in Crs in group 1 and 2. DSPC/TP was significantly lower in group 3,4,5 and 6 than in group 1 ( P 0.05 ) . Conclusion 1.3 MAC isoflurane or sevoflurane mixed with 20 ppm NO can be used safely during mechanical ventilation.