Protective Effect of Ulinastatin on Acute Lung Injury in Diabetic Sepsis Rats

ObjectiveTo establish a rat model of diabetic sepsis, to observe the protective effect of ulinastatin on acute lung injury in diabetic sepsis rats, and to explore the possible mechanism from the aspects of inflammatory response, oxidative stress, hypoxia-inducible factor-1 α expression and pulmonary microvascular permeability. MethodsA total of 50 SPF adult male SD rats were randomly selected, 38 were fed with high fat diet, and the remaining 12 were fed with ordinary diet. Feed five weeks later, the fatty group according to 30 mg/kg body weight, abdominal single injection of STZ, three days after injection, in a random blood glucose or greater tendency for 16.7 L for type 2 diabetes mellitus rats building success, take 36 ChengMo rats were randomly divided into four groups, each group of 12, namely D group of type 2 diabetes, the DS group of type 2 diabetes mellitus and sepsis, Group U ulinastatin pretreatment group. Twelve ordinary feed group were selected as C group, namely blank control group. After successful modeling of type 2 diabetic rats, DS group and U group were injected with endotoxin (LPS) at 5mg/kg via tail vein to construct diabetic sepsis lung injury rat model. Group U was injected with ulinastatin 100kU/kg caudal vein one hour before LPS. 4h later (4 rats in each group were injected with 2 mL 1% Evans blue solution through tail vein half an hour before execution), blood was collected and lung tissues were removed. HE staining was used to observe the pathological changes of lung tissues. The wet/dry ratio (W/D) of lung tissue was determined. Serum IL-1β, IL-18 and TNF-A were detected by ELISA. The contents of malondialdehyde (MDA) and superoxide dismutase (SOD) in serum were detected according to the kit instruction. Western blot was used to detect the hypoxia-inducible factor-1 α (HIF-1α) protein in renal tissue. The expression of TLR4mRNA in lung tissues was detected by rt-pcr. Evans blue staining was used to detect pulmonary microvascular permeability. ResultsCompared with the control group, the lung interstitium in group D was thickened to a certain extent, with a small amount of inflammatory cell infiltration and a little exudate in some alveolar cavities. Compared with D group, DS group had more serious damage, with obvious pulmonary interstitial hyperemia and edema, a large amount of exudation in alveolar cavity, significantly increased inflammatory cells, necrosis and swelling of alveolar epithelial cells. In group U, there were more inflammatory cells in the interstitium and part of alveolar cavities, epithelial cells were exfoliated occasionally in the lumen, and the interstitium widened and the loss was less than that in group DS. Compared with group C, the wet/dry weight ratio (W/D), serum IL-1β, IL-18 and TNF-A contents, hiF-1 α protein expression, TLR4 mRNA expression and pulmonary microvascular permeability of rats in other groups were significantly increased (P<0.01 or P<0.05). The content of MDA and SOD activity in serum decreased (P<0.01). Compared with group D, the wet/dry weight ratio (W/D), serum IL-1β, IL-18 and TNF-A contents, hiF-1 α protein expression, TLR4 mRNA expression and pulmonary microvascular permeability in DS group and U group were significantly increased (P<0.01 or P<0.05). The content of MDA in serum was increased (P<0.01 or P<0.05), and the activity of SOD in DS group was decreased (P<0.01). ; Compared with DS group, the wet/dry weight ratio (W/D), serum IL-1β, IL-18, TNF-A contents, hiF-1 α protein expression, TLR4 mRNA expression, and pulmonary microvascular permeability of rats in GROUP U were significantly decreased (P < 0.01 or P<0.05). The content of MDA in serum decreased (P<0.01), and the activity of SOD increased (P<0.01). ConclusionUlinastatin can effectively reduce acute lung injury induced by diabetic sepsis in rats, and its mechanism may be related to inhibiting inflammatory response, reducing oxidative stress, regulating hypoxia response pathway and improving pulmonary microvascular permeability.

Abstract P808: The Main Peptide Ssubstrates of Neurolysin Enhance Brain Microvascular Permeability in a Human in vitro Model

Increased brain microvascular permeability and disruption of blood-brain barrier (BBB) function are among hallmarks of ischemic stroke. Recently, peptidase neurlysin (Nln) has been identified as a compensatory and cerebroprotective mechanism in the post-stroke brain that functions to process a diverse group of extracellular neuropeptides, including bradykinin (BK), neurotensin (NT) and substance P (SP). A number of studies suggest involvement of BK, NT and SP in BBB impairment and edema formation after stroke, however there is paucity of data in regards to the direct effects of these peptides on the brain microvascular endothelial cells (BMECs) and BBB. The purpose of this study was to evaluate the direct effects of BK, NT and SP on permeability of BBB in an in vitro model based on human, induced pluripotent stem cell (iPSC)-derived BMECs. Our data indicate that all three peptides increase BBB permeability in a concentration-dependent manner in an in vitro model formed from two different iPSC lines (CTR90F and CTR65M) and widely used hCMEC/D3 human BMECs. Combination of BK, NT and SP at a sub-effective concentration also resulted in increased BBB permeability in the iPSC-derived model. Furthermore, we observed abrogation of BK, NT and SP effects with pretreatment of pharmacological blockers targeting their specific receptors or in presence of recombinant neurolysin (Nln). This is the first experimental study to document increased permeability of BBB in response to direct action of NT in an in vitro model. In addition, our study confirms the expected but not well-documented, direct effect of SP on BBB permeability and adds to the well-recognized actions of BK on BBB. Lastly, we demonstrate that peptidase Nln can neutralize the effects of these peptides on BBB, suggesting potential therapeutic implications.

Quantitative Dynamic Contrast-Enhanced Magnetic Resonance Imaging for the Analysis of Microvascular Permeability in Peritumor Brain Edema of Fibrous Meningiomas

<b><i>Introduction:</i></b> This study aims to analyze the permeability of intra- and peri-meningiomas regions and compare the microvascular permeability between peritumoral brain edema (PTBE) and non-PTBE using DCE-MRI. <b><i>Methods:</i></b> This was a retrospective of patients with meningioma who underwent surgery. The patients were grouped as PTBE and non-PTBE. The DCE-MRI quantitative parameters, including volume transfer constant (<i>K</i>^trans), rate constant (<i>K</i>_ep), extracellular volume (<i>V</i>_e), and mean plasma volume (<i>V</i>_p), obtained using the extended Tofts-Kety 2-compartment model. Logistic regression analysis was conducted to explore the risk factor of PTBE. <b><i>Results:</i></b> Sixty-three patients, diagnosed as fibrous meningioma, were included in this study. They were 17 males and 46 females, aged from 32 to 88 years old. <i>K</i>_ep and <i>V</i>_p were significantly lower in patients with PTBE compared with those without (<i>K</i>_ep: 0.1852 ± 0.0369 vs. 0.5087 ± 0.1590, <i>p</i> = 0.010; <i>V</i>_p: 0.0090 ± 0.0020 vs. 0.0521 ± 0.0262, <i>p</i> = 0.007), while there were no differences regarding <i>K</i>^trans and <i>V</i>_e (both <i>p</i> &#x3e; 0.05). The multivariable analysis showed that tumor size ≥10 cm³ (OR = 4.457, 95% CI: 1.322–15.031, <i>p</i> = 0.016) and <i>V</i>_p (OR = 0.572, 95%CI: 0.333–0.981, <i>p</i> = 0.044) were independently associated with PTBE in patients with meningiomas. <b><i>Conclusion:</i></b> DCE-magnetic resonance imaging·Meningioma·Blood vessel MRI can be used to quantify the microvascular permeability of PTBE in patients with meningioma.

Redox Regulation of Microvascular Permeability: IL-1β Potentiation of Bradykinin-Induced Permeability Is Prevented by Simvastatin

Antioxidant effects of statins have been implicated in the reduction in microvascular permeability and edema formation in experimental and clinical studies. Bradykinin (Bk)-induced increases in microvascular permeability are potentiated by IL-1β; however, no studies have examined the protection afforded by statins against microvascular hyperpermeability. We investigated the effects of simvastatin pretreatment on albumin–fluorescein isothiocyanate conjugate (FITC-albumin) permeability in post-capillary venules in rat cremaster muscle. Inhibition of nitric oxide synthase with L-NAME (10µM) increased basal permeability to FITC-albumin, which was abrogated by superoxide dismutase and catalase. Histamine-induced (1 µM) permeability was blocked by L-NAME but unaffected by scavenging reactive oxygen species with superoxide dismutase (SOD) and catalase. In contrast, bradykinin-induced (1–100 nM) permeability increases were unaffected by L-NAME but abrogated by SOD and catalase. Acute superfusion of the cremaster muscle with IL-1β (30 pM, 10 min) resulted in a leftward shift of the bradykinin concentration–response curve. Potentiation by IL-1β of bradykinin-induced microvascular permeability was prevented by the nicotinamide adenine dinucleotide phosphate oxidase (NADPH oxidase) inhibitor apocynin (1 µM). Pretreatment of rats with simvastatin (5 mg·kg−1, i.p.) 24 h before permeability measurements prevented the potentiation of bradykinin permeability responses by IL-1β, which was not reversed by inhibition of heme oxygenase-1 with tin protoporphyrin IX (SnPP). This study highlights a novel mechanism by which simvastatin prevents the potentiation of bradykinin-induced permeability by IL-1β, possibly by targeting the assembly of NADPH oxidase subunits. Our findings highlight the therapeutic potential of statins in the prevention and treatment of patients predisposed to inflammatory diseases.