Dysregulation of p53 and phosphoinositide (PIPn) signaling are both key drivers of oncogenesis and metastasis. Our recent findings reveal a previously unrecognized interaction between these pathways, converging in the nucleus to form a PIPn-p53 signalosome that modulates nuclear AKT activation and downstream signaling, thereby influencing cancer cell survival and motility. This review examines recent insights into nuclear PIPn signaling in the context of established roles for p53 in cell dynamics and migration while also deliberating current research on how nuclear PIPns interact with p53 to form signalosomes that affect cell motility. We emphasize the critical role of PIPns in stabilizing p53 and activating de novo nuclear AKT signaling, which subsequently modulates key motility-related pathways. Understanding the unique operation and function of the PIPn-p53 signalosome in nuclear phosphatidylinositol 3-kinase (PI3K)-AKT activation offers novel therapeutic strategies for controlling cancer metastasis by targeting pertinent interactions and events.
Humans ; Tumor Suppressor Protein p53/metabolism* ; Signal Transduction ; Cell Movement ; Cell Nucleus/metabolism* ; Phosphatidylinositols/metabolism* ; Proto-Oncogene Proteins c-akt/metabolism* ; Animals ; Neoplasms/pathology* ; Phosphatidylinositol 3-Kinases/metabolism*

Objective:To study and verify the molecular mechanism of Duzhong Pills for osteoporosis (OP) by means of network pharmacology and molecular docking.Methods:The main chemical components of Duzhong Pills were mined by TCMSP database and the related targets were predicted. The potential targets of osteoporosis in GeneCards, DisGeNET and OMIM databases were searched and the common targets of both were obtained. The STRING platform was used for protein interaction analysis and PPI network diagram was made. The common targets were imported into the David database for enrichment analysis of GO and KEGG pathways, and molecular docking of the main components and core targets was performed. Eighteen Sprague-Dawley rats were divided into control group, model group and Duzhong Pills group according to random number table method, with 6 rats in each group. Ovariectomy was used to make osteoporosis model in model group and Duzhong Pills group. Duzhong Pills group was intragaxed with Duzhong Pills extract of 5 g/kg, and control group and model group were intragaxed with normal saline of the same volume, once a day for 8 weeks. Serum levels of TNF-α, IL-6 and IL-1β were detected by ELISA, and femur PI3K and Akt were detected by Western blot.Results:34 active components were obtained from Duzhong Pills, corresponding to 243 targets, and 1 059 targets for osteoporosis. The core targets included TNF-α, IL-6, AKT1, TP53, IL-1β and others regulated osteoporosis through PI3K-Akt and TNF pathway. The experimental results indicated that compared with model group, the levels of serum TNF-α, IL-6 and IL-1β in Duzhong Pills group decreased ( P<0.05), and the expressions of PI3K and Akt in femur decreased ( P<0.05). Conclusion:Through β-sitosterol, quercetin, kaempferol and other active components, Duzhong Pills can act on TNF, IL-6, AKT1, TP53, IL-1β and other targets, regulating PI3K-Akt signaling pathway, TNF signaling pathway and other signaling pathways to play a role in the treatment of osteoporosis.

Objective:To explore the protective effect of sivelestat (SV) against sepsis-induced acute kidney injury (AKI) and its molecular mechanism.Methods:According to the random number table method, 64 male Wistar rats were divided into sham operation group (Sham group), sepsis due to cecal ligation and puncture group (CLP group), low dose of SV treatment group (SL group, 50 mg/kg SV was injected into the tail vein at 12 hours and 24 hours after CLP), and high dose of SV treatment group (SH group, 100 mg/kg SV was injected into the tail vein at 12 hours and 24 hours after CLP), with 16 rats in each group. 48 hours after CLP, the 48-hour survival of rats were recorded, all rats were sacrificed and samples were harvested. Enzyme-linked immunosorbent assay (ELISA) was used to detect the serum levels of kidney injury molecule-1 (KIM-1), interleukins (IL-1β, IL-6), tumor necrosis factor-α (TNF-α) and neutrophil elastase (NE). Hematoxylin-eosin (HE) staining was used to observe histopathological changes and assess renal tubule injury score. Masson staining was used to detect the collagen volume fraction (CVF) of kidney tissue. Western blotting was used to detect the protein expressions of phosphatidylinositol 3-kinase (PI3K), phosphorylation PI3K (p-PI3K), protein kinase B (AKT), phosphorylation AKT (p-AKT), nuclear factor-κB p65 (NF-κB p65) and NE. The protein expressions of p-PI3K, p-AKT, NF-κB p65 were detected by immunohistochemistry.Results:Compared with Sham group, the 48-hour survival rate of CLP group was significantly reduced. Histopathological results showed that large tubular epithelial cells and brush margins were shed, tubular casts were formed, some tubular atrophy, glomerular hyperemia, renal interstitial inflammatory cell infiltration and increased renal tubular injury score. Renal interstitial fibrosis was obvious and CVF increased. The levels of KIM-1, IL-1β, IL-6, TNF-α and NE in serum were significantly elevated in the CLP group. The proteins expression of inflammatory pathway-related p-PI3K/PI3K, p-AKT/AKT, NF-κB p65 and NE were significantly increased in kidney tissue. It suggested that septic rats had renal injury and the PI3K/AKT inflammatory pathway was activated. Compared with CLP group, there was no significant difference in 48-hour survival in SL group and SH group (68.75%, 75.00% vs. 56.25%, both P > 0.05), but kidney injury was significantly relieved. Specifically: renal tubular injury score and CVF significantly decreased [tubular injury score: 2 (1, 2), 1 (1, 1) vs. 2 (2, 3); CVF: (22.36±0.86)%, (18.74±1.05)% vs. (58.38±0.79)%, all P < 0.05]; the serum levels of KIM-1, IL-1β, IL-6, TNF-α and NE also decreased significantly [KIM-1 (ng/L): 145.03±8.88, 117.58±7.02 vs. 158.22±12.00; IL-1β (ng/L): 108.32±9.00, 92.98±8.06 vs. 133.78±8.48; IL-6 (ng/L): 124.33±10.11, 115.42±8.17 vs. 165.19±5.70; TNF-α (ng/L): 321.56±19.29, 289.68±21.57 vs. 424.88±22.76, NE (mol/L): 93.84±9.14, 75.01±10.56 vs. 113.45±6.39, all P < 0.05]; the proteins expression of inflammatory pathway-related p-PI3K/PI3K, p-AKT/AKT, NF-κB p65 and NE were significantly decreased (p-PI3K/PI3K: 0.93±0.06, 0.67±0.04 vs. 1.27±0.08; p-AKT/AKT: 0.78±0.09, 0.47±0.05 vs. 0.96±0.12; NF-κB p65/GAPDH: 1.43±0.13, 0.85±0.08 vs. 1.88±0.17; NE/GAPDH: 1.45±0.06, 0.91±0.04 vs. 1.71±0.08, all P < 0.05), the positive expressions of p-PI3K, p-AKT and NF-κB p65 in kidney tissue were decreased [p-PI3K positive expression area: (13.36±1.84)%, (8.03±1.12)% vs. (21.56±1.20)%; p-AKT positive expression area: (21.57±0.91)%, (15.21±2.76)% vs. (30.81±2.12)%; NF-κB p65 positive expression area: (25.17±1.38)%, (17.07±2.11)% vs. (37.85±2.50)%, all P < 0.05]. Serum inflammatory factor level, and PI3K/AKT pathway related protein, NF-κB p65, NE protein expression level and p-PI3K, p-AKT, NF-κB p65 positive area and other indicators in renal tissue in SH group were further lower than those in SL group (all P < 0.05). Conclusions:SV can ameliorate sepsis-induced AKI. The mechanism may be related to the inhibition of PI3K/AKT pathway, and high dose of SV has better efficacy.

Objective:To investigate the protective effect of β- blocker (esmolol) on myocardia and toll-like receptor 4 (TLR4) inflammatory pathway in septic rats.Methods:Sixty male Wistar rats were randomly (random number) divided into the shame group, sepsis group (CLP group), esmolol group (CLP+ES group) and TLR4 inhibitor group (CLP+TAK-242 group) with 15 rats in each group. Cecal exploration was performed in the shame group, and cecal ligation and perforation (CLP) was performed in the CLP group, CLP+ES group and CLP+TAK-242 group. The CLP+ES group received intraperitoneal injection of esmolol diluent 20 mg/kg 12 h after CLP. The CLP+TAK-242 group was given intraperitoneal injection of TAK-242 3 mg/kg at the same time point as above. The shame group and CLP group were given the same amount of normal saline. Rats in all groups were sacrificed 24 h after operation, and the samples were collected and processed. The pathological changes of myocardium were observed by hematoxylin - eosin staining. The expression of TLR4, myeloid differentiation protein 88 (MyD88) and nuclear factor -κB (NF-κB) in myocardial tissue were observed by immunohistochemistry. Masson staining was used to observe the expression of fibers and inflammatory factors in myocardial tissue. The protein expressions of TLR4, MyD88, NF-κB and aspartic acid specific cysteine protease 1 (caspase-1) were detected by Western blot. Serum levels of cardiac troponin I (cTn-I), tumor necrosis factor α (TNF-α), interleukin-6 (IL-6) and interleukin-1β (IL-1β) were detected by enzyme-linked immunosorbent assay (ELISA).Results:Compared with the shame group, myocardial injury, fibrosis and inflammatory cell infiltration were significantly aggravated in the CLP group, and the levels of myocardial injury index cTn-I and inflammatory mediators TNF-α, IL-6 and IL-1β were significantly increased [(8.70±0.22) vs. (4.41±0.31), (445.57±9.13) vs. (219.60±5.52), (165.55±2.18) vs. (93.47±3.37), (124.12±2.59) vs. (67.63±6.04),all P<0.05]. Compared with the CLP group, myocardial injury was significantly reduced in the CLP+ES group and CLP+TAK-242 group, and the levels of inflammatory transmitters were significantly reduced [(5.38±0.18) and (5.37±0.13) vs. (8.70±0.22), (322.73±7.63) and (300.58±17.47) vs. (445.57±9.13), (121.28±5.44) and (120.30±4.95) vs. (165.55±2.18), (102.60±4.09) and (105.08±7.21) vs. (124.12±2.59), all P<0.05]. Western blot analysis showed that the protein expression levels of TLR4, MyD88, NF-κB and caspase-1 in the CLP group were significantly higher than those in the shame group [(1.79±0.15) vs. (1.15±0.04), (4.70±0.30) vs. (3.87±0.10), (0.35±0.04) vs. (0.18±0.02), (2.27±0.29) vs. (1.15±0.07), all P<0.05], while the protein expression levels in the CLP+ES group and CLP+TAK-242 group were significantly lower than those in the CLP group [(1.31±0.16) and (1.18±0.14) vs. (1.79±0.15), (1.50±0.16) and (1.46±0.19) vs. (2.27±0.29), (0.27±0.02) and (0.24±0.01) vs. (0.35±0.04), (1.50±0.16) and (1.46±0.19) vs. (2.27±0.29), all P<0.05]. Conclusions:β-blocker can reduce myocardial injury and inhibit the expression of inflammatory mediators in septic rats by blocking the inflammatory response mediated by TLR4 signaling pathway.

Objective:To explore the basic characteristics of various types of intensive care unit (ICU) patients and the predictive value of six common disease severity scores in critically ill patients on the first day on the 28-day death risk.Methods:The general information, disease severity scores [acute physiology score Ⅲ (APSⅢ), Oxford acute disease severity (OASIS) score, Logistic organ dysfunction score (LODS), simplified acute physiology score Ⅱ (SAPSⅡ), systemic inflammatory response syndrome (SIRS) score and sequential organ failure assessment (SOFA) score], prognosis and other indicators of critically ill patients admitted from 2008 to 2019 were extracted from Medical Information Mart for Intensive Care-Ⅳ 2.0 (MIMIC-Ⅳ 2.0). The receiver operator characteristic curve (ROC curve) of six critical illness scores for 28-day death risk of patients in various ICU, and the area under the ROC curve (AUC) was calculated, the optimal Youden index was used to determine the cut-off value, and the AUC of various ICU was verified by Delong method.Results:A total of 53 150 critically ill patients were enrolled, with medical ICU (MICU) accounted for the most (19.25%, n = 10 233), followed by cardiac vascular ICU (CVICU) with 17.78% ( n = 9 450), and neurological ICU (NICU) accounted for the least (6.25%, n = 3 320). The patients in coronary care unit (CCU) were the oldest [years old: 71.79 (60.27, 82.33)]. The length of ICU stay in NICU was the longest [days: 2.84 (1.51, 5.49)] and accounted for the highest proportion of total length of hospital stay [63.51% (34.61%, 97.07%)]. The patients in comprehensive ICU had the shortest length of ICU stay [days: 1.75 (0.99, 3.05)]. The patients in CVICU had the lowest proportion of length of ICU stay to total length of hospital stay [27.69% (18.68%, 45.18%)]. The six scores within the first day of ICU admission in NICU patients were lower than those in the other ICU, while APSⅢ, LODS, OASIS, and SOFA scores in MICU patients were higher than those in the other ICU. SAPⅡ and SIRS scores were both the highest in CVICU, respectively. In terms of prognosis, MICU patients had the highest 28-day mortality (14.14%, 1 447/10 233), while CVICU patients had the lowest (2.88%, 272/9 450). ROC curve analysis of the predictive value of each score on the 28-day death risk of various ICU patients showed that, the predictive value of APSⅢ, LODS, and SAPSⅡ in comprehensive ICU were higher [AUC and 95% confidence interval (95% CI) were 0.84 (0.83-0.85), 0.82 (0.81-0.84), and 0.83 (0.82-0.84), respectively]. The predictive value of OASIS, LODS, and SAPSⅡ in surgical ICU (SICU) were higher [AUC and 95% CI were 0.80 (0.79-0.82), 0.79 (0.78-0.81), and 0.79 (0.77-0.80), respectively]. The predictive value of APSⅢ and SAPSⅡ in MICU were higher [AUC and 95% CI were 0.84 (0.82-0.85) and 0.82 (0.81-0.83), respectively]. The predictive value of APSⅢ and SAPSⅡ in CCU were higher [AUC and 95% CI were 0.86 (0.85-0.88) and 0.85 (0.83-0.86), respectively]. The predictive value of LODS and SAPSⅡ in trauma ICU (TICU) were higher [AUC and 95% CI were 0.83 (0.82-0.83) and 0.83 (0.82-0.84), respectively]. The predictive value of OASIS and SAPSⅡ in NICU were higher [AUC and 95% CI were 0.83 (0.80-0.85) and 0.81 (0.78-0.83), respectively]. The predictive value of APSⅢ, LODS, and SAPSⅡ in CVICU were higher [AUC and 95% CI were 0.84 (0.83-0.85), 0.81 (0.80-0.82), and 0.78 (0.77-0.78), respectively]. Conclusions:For the patients in comprehensive ICU, MICU, CCU, and CVICU, APSⅢ or SAPSⅡ can be applied for predicting 28-day death risk. For the patients in SICU and NICU, OASIS or SAPSⅡ can be applied to predict 28-day death risk. For the patients in TICU, SAPSⅡ or LODS can be applied for predicting 28-day death risk. For CVICU patients, APSⅢ or LODS can be applied to predict 28-day death risk.

Sepsis is a life-threatening organ dysfunction caused by dysregulation of the body's response to infection. It is one of the common and serious complications in clinically critical patients with trauma, burn, shock, infection, etc., with high morbidity and mortality. Although the treatment of sepsis has made great achievements in clinical practice, the mortality of patients with sepsis is still increasing due to its secondary complications. Septic cardiomyopathy (SCM) is one of the major complications that threaten septic patient's life. SCM refers to myocardial dysfunction with the aggravation of the primary disease, which is manifested by biventricular dilatation accompanied by a decrease in left ventricular ejection fraction (LVEF). It is one of the major complications that threaten the life of patients with sepsis. The existing research shows that the mechanism of SCM includes myocardial mitochondrial dysfunction, myocardial cell apoptosis, calcium circulation disorder and its treatment including conventional treatment, β 1 receptor blocker treatment and traditional Chinese medicine treatment, etc. This paper reviewed the pathogenesis of SCM and its related, in order to provide references for the rational diagnosis and treatment of SCM.

Objective:To explore the damage of the intestinal mucosal barrier of septic rats by the activation of NOD-like receptor family, pyrin domain-containing 3 (NLRP3) inflammasomes and the role of Ulinastatin (UTI) on the expression of intestinal nuclear factor-κB (NF-κB)/NLRP3 inflammasome signaling pathway in septic rats.Methods:According to the random number table method, 64 male Wistar rats were divided into sham operation group (Sham group), cecal ligation and puncture (CLP) group, UTI treatment group (100 kU/kg UTI was intraperitoneally injected 1, 6, 12 and 18 hours after CLP), and UTI pretreatment group (100 kU/kg UTI was given 1 hour before CLP), with 16 rats in each group. The survival of rats was observed after 24 hours, and the blood was collected from abdominal aorta at 24 hours after modeling, then rats were killed and their ileum tissues were taken. Hematoxylin-eosin (HE) staining was used to observe histopathological changes and Chiu score. The levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and intestinal fatty acid binding protein (I-FABP) in serum were detected by enzyme linked immunosorbent assay (ELISA). The protein expression of NF-κB p65 in intestinal tissue was detected by Western blotting. The expression of intestinal tight junction proteins Claudin-1, Occludin and the inflammasome NLRP3, apoptosis-associated speck-like protein containing CARD (ASC) and caspase-1 were detected by immunohistochemistry.Results:Compared with Sham group, the 24-hour survival rate of CLP group was significantly reduced. Histopathological results showed that the CLP group had severe edema of mucosa and submucosal stroma with obvious infiltration of inflammatory cells and disordered villi arrangement. Some glands were incomplete, and the villus structure was severely damaged. The Chiu score was significantly increased. The levels of TNF-α, IL-1β, I-FABP in serum and the protein expression of NF-κB p65 in intestinal tissue were significantly increased. The positive expressions of NLRP3, caspase-1 and ASC were also significantly increased. However, the positive expression of tight junction protein in small intestine tissue such as Occludin and Claudin-1 were significantly reduced. It suggested that when sepsis occurs, small intestinal mucosal barrier dysfunction happens, and mucosal permeability increases, while tight junction protein expression decreases, NLRP3 inflammasome and its upstream molecule NF-κB p65 were activated. After UTI treatment and UTI pretreatment intervention, although there was no significant difference in 24-hour survival compared with CLP group (62.5%, 68.8% vs. 43.8%, both P > 0.05), the intestinal tissue damage of septic rats was significantly improved. Specifically: Chiu score and the levels of TNF-α, IL-1β, I-FABP in serum were significantly decreased [Chiu score: 3.37±0.25, 3.23±0.16 vs. 4.08±0.13, TNF-α (ng/L): 147.62±20.74, 140.71±24.81 vs. 222.82±16.84, IL-1β (ng/L): 80.64±5.68, 78.11±4.75 vs. 133.73±3.92, I-FABP (μg/L): 38.29±3.60, 35.88±4.52 vs. 59.81±4.66, all P < 0.05]; the protein expression of NF-κB p65 was significantly decreased (NF-κB p65/β-actin: 0.65±0.10, 0.69±0.11 vs. 0.99±0.10, both P < 0.05), the positive expressions of Claudin-1 and Occludin in the small intestine tissue were increased [Claudin-1 positive expression area: (19.43±3.08)%, (23.99±6.27)% vs. (7.77±2.03)%; Occludin positive expression area: (19.58±4.75)%, (23.28±3.68)% vs. (11.69±4.30)%, all P < 0.05], while the positive expressions of NLRP3, caspase-1, ASC were decreased [NLRP3 positive expression area: (7.80±3.14)%, (6.86±2.63)% vs. (14.44±3.68)%; caspase-1 positive expression area: (10.62±3.52)%, (9.49±3.09)% vs. (26.69±8.05)%; ASC positive expression area: (9.95±2.81)%, (10.53±3.61)% vs. (24.16±5.48)%, all P < 0.05]. However, there was no significant difference in the improvement effect between UTI treatment group and UTI pretreatment group.Conclusions:Intestinal barrier dysfunction in sepsis may be related to the activation of NLRP3 inflammasomes in the intestinal mucosa. The protective effect of UTI in the intestinal mucosa may be related to inhibiting the activation of NLRP3 inflammasomes in the intestinal mucosa, but UTI pretreatment has no obvious advantage compared with UTI treatment.

Objective:To explore whether resveratrol (RSV) could activate silent information regulator 1 (SIRT1) to regulate the activation of NOD-like receptor protein 3 (NLRP3) inflammasome in sepsis induced intestinal injury model, and then reduce intestinal inflammation and cell apoptosis, so as to play a protective role in intestinal barrier function.Methods:① In vitro experiment: human Colorectal adenocarcinoma cells (Caco-2) were cultured, which were divided into normal group (normal culture on complete medium for 48 hours), lipopolysaccharide (LPS) group (normal culture on complete medium for 24 hours, then LPS containing 2 mg/L complete medium intervention for 6 hours), RSV low, medium and high concentration groups and SIRT1 inhibitor (EX-527) group (complete medium normal culture for 24 hours, LPS containing 2 mg/L complete medium intervention for 6 hours, followed by RSV 10, 20, 40 μmol/L or EX-527 10 μmol/L intervention for 6 hours, respectively). The levels of tumor necrosis factor-α (TNF-α) and interleukins (IL-6, IL-18, IL-1β) in the cell supernatant were determined by enzyme linked immunosorbent assay (ELISA). The apoptosis level of the cells was detected by flow cytometry. Western blotting was used to detect the protein levels of NLRP3, SIRT1, caspase-1 and apoptosis-associated speck-like protein containing a CARD (ASC). ② In vivo experiment: according to random number table method, 24 male Wistar rats were divided into sham operation group (Sham group), cecal ligation and perforation (CLP) 6 hours group (CLP 6 h group), CLP 24 h group and RSV intervention group [RSV (20 mg/kg) was intraperitoneally injected 6 hours and 12 hours after CLP], with 6 rats in each group. The levels of NLRP3, caspase-1 and ASC in the intestine of rats were detected by immunohistochemistry. Results:① Compared with the normal group, the levels of inflammatory factors in the cell supernatant of the LPS group were increased and the expression of SIRT1 protein was decreased, while the protein expressions of NLRP3, caspase-1 and ASC were increased. Compared with LPS group, different concentrations of RSV reduced the level of inflammatory factors, increased the activity of SIRT1, inhibited the expression of NLRP3 inflammasome and its downstream products caspase-1 and ASC, and the effect of high concentration of RSV (40 μmol/L) was the most significant [TNF-α (ng/L): 8.77±0.43 vs. 12.66±0.81, IL-6 (ng/L): 1.35±0.20 vs. 1.93±0.09, IL-1β (ng/L): 1.05±0.04 vs. 1.31±0.07, IL-18 (ng/L): 519.50±11.16 vs. 622.70±30.69, SIRT1/β-actin: 0.80±0.05 vs. 0.58±0.02, caspase-1/β-actin: 0.55±0.06 vs. 0.78±0.06, ASC/β-actin: 0.78±0.08 vs. 1.04±0.15, all P < 0.05], while SIRT1 inhibitor EX-527 had the opposite effects. There was no significant difference in the apoptosis rate among normal group, LPS group, and low, medium and high concentration RSV groups, as well as EX-527 group [(7.03±0.57)%, (9.67±0.55)%, (9.57±0.70)%, (9.30±2.15)%, (9.87±0.97)%, (9.07±0.93)%, F = 2.590, P = 0.082]. ② Immunohistochemical results showed that compared with the Sham group, the expressions of NLRP3 inflammasomes and downstream products caspase-1 and ASC in the intestinal epithelial cells in CLP 6 h group, CLP 24 h group and RSV intervention group were significantly increased. The percentage of ASC-positive area in intestinal epithelium of RSV intervention group was significantly lower than that of CLP 6 h group [(15.22±2.73)% vs. (19.88±2.67)%, P < 0.05], and the expressions of NLRP3 and caspase-1 were significantly lower than those of CLP 24 h group [(9.31±1.37)% vs. (13.19±1.92)%, (19.57±3.92)% vs. (27.28±6.33)%, both P < 0.05]. Conclusion:After sepsis, high concentration of RSV could inhibit the activation of NLRP3 inflammasome by activating SIRT1, thereby reduce the expression of caspase-1 and ASC, and inhibit the secretion of inflammatory factors to reduce the inflammatory response.

Objective:To investigate the expression of NOD-like receptor protein 3 (NLRP3) inflammasome in intestinal injury models with different severity of sepsis and the inflammatory response and apoptosis mediated by NLRP3 inflammasome.Methods:Human colorectal adenocarcinoma cells (Caco-2) were cultured in vitro. The logarithmic growth phase cells were divided into blank control group (normal culture in complete medium) and lipopolysaccharide (LPS) 1, 2 and 4 mg/L groups (complete medium containing 1, 2 and 4 mg/L LPS, respectively). The supernatant were collected at 6, 12 and 24 hours, and the levels of tumor necrosis factor-α (TNF-α), interleukins (IL-6, IL-1β, IL-18) were detected by enzyme linked immunosorbent assay (ELISA). The apoptotic level of cells was detected by flow cytometry. The cells were harvested, and the real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to detect the mRNA expressions of NLRP3 and silent information regulator 1 (SIRT1). Western blotting was used to detect the protein expressions of NLRP3, SIRT1, caspase-1 and apoptosis-associated speck-like protein (ASC). Results:ELISA results showed that the levels of IL-6, TNF-α, IL-1β, and IL-18 in cell supernatant of LPS groups increased in a dose-dependent and time-dependent manner as compared with the blank control group during the same intervention period. The increase was most significant in LPS 4 mg/L group at 24 hours [IL-6 (ng/L): 3.55±0.06 vs. 0.67±0.09, TNF-α (ng/L): 15.37±0.19 vs. 5.04±0.14, IL-1β (ng/L): 2.26±0.10 vs. 0.56±0.09, IL-18 (ng/L): 433.92±22.55 vs. 93.55±21.13, all P < 0.05]. The results of the apoptotic test showed that, compared with the blank control group, the apoptotic rate of LPS groups increased in a dose-dependent and time-dependent manner, and the apoptotic rate of LPS 4 mg/L group increased most significantly at 24 hours [(14.83±3.73)% vs. (5.87±1.17)%, P < 0.05]. RT-qPCR results showed that the expression level of NLRP3 mRNA was increased, while the expression level of SIRT1 mRNA was decreased with the increase of LPS intervention dose and the prolonging of intervention time. At 24 hours, there were significant differences between LPS 4 mg/L group and blank control group [NLRP3 mRNA (2 -ΔΔCt): 8.20±2.82 vs. 1.00±0.36, SIRT1 mRNA (2 -ΔΔCt): 0.58±0.01 vs. 1.03±0.06, both P < 0.05]. Western blotting showed that compared with the blank control group, the protein expression levels of NLRP3, caspase-1 and ASC in LPS groups were significantly increased, while the protein expression levels of SIRT1 were significantly decreased. During each intervention period, with the increase of LPS dose, the expressions of NLRP3, caspase-1 and ASC protein increased gradually, while the expression of SIRT1 protein decreased gradually. At 24 hours, the difference between the LPS 4 mg/L group and the blank control group was significant [NLRP3 protein (NLRP3/β-actin): 1.48±0.03 vs. 0.90±0.12, caspase-1 protein (caspase-1/β-actin): 1.18±0.11 vs. 0.72±0.09, ASC protein (ASC/β-actin) : 1.09±0.01 vs. 0.82±0.03, SIRT1 protein (SIRT1/β-actin): 0.48±0.03 vs. 0.76±0.05, all P < 0.05]. Conclusion:In vitro, in the sepsis induced intestinal inflammation model, with the increase of LPS intervention dose and the prolongation of intervention time, intestinal inflammatory response and cell apoptosis showed an increasing trend, which may be related to the up-regulation of NLRP3 inflammasome and its downstream products ASC and caspase-1, and to the down-regulation of SIRT1 expression.

Objective:To summarize the patient profiles and therapeutic efficacies of ABO-incompatible living-related kidney transplantations at 19 domestic transplant centers and provide rationales for clinical application of ABOi-KT.Methods:Clinical cases of ABO-incompatible/compatible kidney transplantation (ABOi-KT/ABOc-KT) from December 2006 to December 2009 were collected. Then, statistical analyses were conducted from the aspects of tissue matching, perioperative managements, complications and survival rates of renal allograft or recipients.Results:Clinical data of 342 ABOi-KT and 779 ABOc-KT indicated that (1) no inter-group differences existed in age, body mass index (BMI), donor-recipient relationship or waiting time of pre-operative dialysis; (2) ABO blood type: blood type O recipients had the longest waiting list and transplantations from blood type A to blood type O accounted for the largest proportion; (3) HLA matching: no statistical significance existed in mismatch rate or positive rate of PRA I/II between two types of surgery; (4) CD20 should be properly used on the basis of different phrases; (5) hemorrhage was a common complication during an early postoperative period and microthrombosis appeared later; (6) no difference existed in postoperative incidence of complications or survival rate of renal allograft and recipients at 1/3/5/10 years between ABOi-KT and ABOc-KT. The acute rejection rate and serum creatinine levels of ABOi-KT recipients were comparable to those of ABOc-KT recipients within 1 year.Conclusions:ABOi-KT is both safe and effective so that it may be applied at all transplant centers as needed.