Development and external validation of a nomogram for overall survival after curative resection in serosa-negative, locally advanced gastric cancer.

BACKGROUND: Few nomograms can predict overall survival (OS) after curative resection of advanced gastric cancer (AGC), and these nomograms were developed using data from only a few large centers over a long time period. The aim of this study was to develop and externally validate an elaborative nomogram that predicts 5-year OS after curative resection for serosa-negative, locally AGC using a large amount of data from multiple centers in Japan over a short time period (2001-2003). PATIENTS AND METHODS: Of 39 859 patients who underwent surgery for gastric cancer between 2001 and 2003 at multiple centers in Japan, we retrospectively analyzed 5196 patients with serosa-negative AGC who underwent Resection A according to the 13th Japanese Classification of Gastric Carcinoma. The data of 3085 patients who underwent surgery from 2001 to 2002 were used as a training set for the construction of a nomogram and Web software. The data of 2111 patients who underwent surgery in 2003 were used as an external validation set. RESULTS: Age at operation, gender, tumor size and location, macroscopic type, histological type, depth of invasion, number of positive and examined lymph nodes, and lymphovascular invasion, but not the extent of lymphadenectomy, were associated with OS. Discrimination of the developed nomogram was superior to that of the TNM classification (concordance indices of 0.68 versus 0.61; P < 0.001). Moreover, calibration was accurate. CONCLUSIONS: We have developed and externally validated an elaborative nomogram that predicts the 5-year OS of postoperative serosa-negative AGC. This nomogram would be helpful in the assessment of individual risks and in the consideration of additional therapy in clinical practice, and we have created freely available Web software to more easily and quickly predict OS and to draw a survival curve for these purposes.

A phase I study evaluating tolerability, pharmacokinetics, and preliminary efficacy of L-menthol in upper gastrointestinal endoscopy.

Peppermint oil has been shown to relax gastrointestinal smooth muscle. In this randomized, placebo-controlled study, an L-menthol preparation, NPO-11, was assessed for tolerability and pharmacokinetics (PK) during gastrointestinal endoscopy. Single doses of NPO-11, as high as 320 mg, were well tolerated. NPO-11 was rapidly absorbed, with peak concentrations reached within 1 h after administration. Approximately 70% of the administered L-menthol and its metabolites were excreted in the urine, and this amount fluctuated with no change in the dose. The principal metabolite identified in plasma and urine was menthol glucuronide. The other metabolites include mono- or di-hydroxylated menthol derivatives, most of which are excreted, in part, as glucuronic acid conjugates. The pharmacokinetic data indicated that when NPO-11 is sprayed directly onto the gastric mucosa, it is rapidly metabolized to glucuronic acid conjugates that are excreted in urine. The findings from this study provide new data on the safety and PK of NPO-11 and support further trials.

Unique characteristics of radiation-induced apoptosis in the postnatally developing small intestine and colon of mice.

Abstract We examined the response of the developing mouse intestine to X radiation using neonates (1 day postpartum), infants (2 weeks postpartum) and adults (7 weeks postpartum). Irradiated adult small intestinal crypts displayed two waves of apoptosis. The first wave peaked at 3 h and was followed by a broad wave with a peak persisting from 24 to 48 h. p53 was expressed during the first wave but not the second wave. For the infant small intestine, the intensity of the first wave was approximately half that of the adult wave, and for the colon the intensity was even smaller. In neonates, apoptosis was delayed, peaking at 6 h for small intestinal crypts and at 24 h for colonic crypts. Although no apoptosis occurred at 3 h postirradiation in neonates, p53 was present in both the small intestine and colon, owing at least in part to the inability of p53 to increase the level of Noxa, a p53-dependent pro-apoptosis protein, suggesting a discontinuity in the p53-Noxa-caspase pathway in neonates. By contrast, the induction of p21, a pro-survival protein, was greater in neonatal cells than in adult cells. Thus it appears that the developing and adult intestine mount distinct apoptotic responses to radiation.

Endoscopic submucosal dissection as a staging measure may not lead to worse prognosis in early gastric cancer patients with additional gastrectomy.

BACKGROUND: Endoscopic submucosal dissection is a novel endoluminal technique that enables resection of early stage gastrointestinal malignancies in an en bloc fashion. AIM: To assess whether preceding endoscopic submucosal dissection affected the prognoses of patients who underwent additional gastrectomy with lymph node dissection due to suspicion of nodal metastasis from endoscopic submucosal dissection specimens. PATIENTS AND METHODS: Thirty-one patients with early gastric cancer who underwent gastrectomy after endoscopic submucosal dissection were retrospectively investigated in terms of their survival and tumour recurrence. Additional gastrectomy was performed when histology of the endoscopic submucosal dissection specimens revealed that the tumours did not meet the criteria for node-negative cancers. RESULTS: Twenty-three (74%) and eight (26%) patients had undergone endoscopic submucosal dissection previously due to clinical diagnoses of node-negative cancers and possible node-positive cancers, respectively. Histology of the resected stomachs and lymph nodes revealed residual carcinoma of the stomach in two (6.5%) patients and nodal metastases in four (13%) patients. All patients remain alive without recurrence (median follow-up, 3.4 years; range, 0.6-5.2 years). CONCLUSIONS: Based on the histology of endoscopic submucosal dissection specimens, preceding endoscopic submucosal dissection itself had no negative influence on a patient's prognosis when additional gastrectomy was performed. It may be permissible to resect some early gastric cancers by endoscopic submucosal dissection as a first step to prevent unnecessary gastrectomy, if technically resectable.

A single institutional non-randomized retrospective comparison between definitive chemoradiotherapy and radical surgery in 82 Japanese patients with resectable esophageal squamous cell carcinoma.

This retrospective study was conducted to compare the treatment results between radical surgery and definitive chemoradiotherapy for resectable squamous cell carcinoma of the esophagus. Between June 2000 and May 2005, 82 consecutive patients were selected for this study in which 33 were treated with chemoradiotherapy and 49 with surgery. The patients in the chemoradiotherapy (CRT) group received 2-4 cycles of 5-fluorouracil (1000 mg/m(2)/day, day 1-4, continuous) combined with cisplatin (75 mg/m(2), day 1, bolus) plus 50.4 Gy of radiation, while those in the surgery group were treated by an esophagectomy with radical node dissection. Eighteen surgical patients received postoperative chemotherapy. The baseline clinical TNM stage was similar between the two groups. With a median follow-up period of 36 months (range: 23-84 months) with 47 survivors (57%), the 3-year overall survival rates (P = 0.22) and disease-free survival rates (P = 0.16) were 48% and 44% in the chemoradiotherapy group versus 65% and 59% in the surgery group, and lacked statistical significance. This non-randomized study on patients with resectable squamous cell carcinoma of the esophagus showed that chemoradiotherapy could result in survival comparable with conventional surgery in spite of selection bias of patients. There is a trend toward improved survival with surgery versus definitive CRT.

Ex vivo pilot study using computed analysis of endo-cytoscopic images to differentiate normal and malignant squamous cell epithelia in the oesophagus.

BACKGROUND AND STUDY AIMS: An endo-cytoscopy system allows acquisition of optical biopsies that are quite similar to conventional histology. To simplify discrimination between normal and malignant tissue in the oesophagus using endo-cytoscopy system, we analysed the nuclear (dark staining) area in the obtained images with the goal of an accurate, automatic diagnosis. PATIENTS AND METHODS: Ex vivo endo-cytoscopic observation was performed using endoscopically or surgically resected oesophagus from 10 enrolled patients. Oesophageal tissues were stained using 1% methylene blue, and endo-cytoscopic images were obtained at normal and malignant areas (two areas of each) in each oesophagus. The centre of each image (4x10(-2) mm(2)) was processed by computer, and the area occupied by the total nuclei in each selected field and its ratio to the entire field were calculated. RESULTS: The mean area of the total nuclei was 0.10x10(-2)+/-0.03x10(-2) mm(2) (range 0.05x10(-2) to 0.18x10(-2) mm(2)) in the normal group and 0.40x10(-2)+/-0.06x10(-2) mm(2) (range 0.33x10(-2) to 0.55x10(-2) mm(2)) in the malignant group (P<0.001). The mean ratio of total nuclei to the entire selected field was 6.4+/-1.9% (range 3.1-11.3%) in the normal tissues and 25.3+/-3.8% (range 20.5-34.5%) in the malignant samples (P<0.001). CONCLUSIONS: Endo-cytoscopy system allowed automatic differentiation of normal and malignant tissues in the oesophagus, which could simplify endo-cytoscopic diagnosis. Further study will elucidate whether such analysis is applicable to inflammatory or pre-malignant epithelia in the oesophagus or other gastrointestinal organs.

Gastric and intestinal phenotypic correlation between exocrine and endocrine components in human stomach tumors.

We have previously suggested that an origin of a stomach cancer is from a progenitor cell specializing toward exocrine cell (Exo-cell) lineages. To clarify whether our hypothesis is correct or not, we analyzed the expression of Exo-cell and endocrine cell (End-cell) markers in a series of lesions for comparison. We evaluated chromogranin A (CgA) expression in 37 early and 73 advanced stomach cancers, in 30 stomach adenomas, in 8 carcinoid tumors, and in 4 endocrine cell carcinomas (ECCs) with assessment of gastric and/or intestinal (G/I) phenotypes in both Exo-cell and End-cell by immunohistochemistry. CgA expression was observed in 10.8% of the early and 16.4% of the advanced stomach cancers, respectively. The End-cell G/I phenotypes were in line with the Exo-cell counterparts in the CgA-positive stomach cancerous areas, and there was strong association between Cdx2 expression and the intestinal End-cell markers. All of the adenoma cases had the intestinal Exo-cell phenotypic expression, with the positive link between Exo-cell and End-cell G/I phenotypes. All stomach carcinoids had CgA expression but no expression of Exo-cell markers. In conclusion, most stomach cancers might develop from a progenitor cell specializing towards Exo-cell lineages, but some cases possessed both Exo-cell and End-cell markers with maturely differentiated phenotypes. In such cases, Exo-cell and End-cell phenotypes were found to correlate strongly, suggesting the possibility of histogenesis from "cancer stem cells".

Differential expression of peroxisome proliferator-activated receptor in histologically different human gastric cancer tissues.

Gastric cancer cell lines express peroxisome proliferator-activated receptor gamma (PPARgamma), and treatment with PPARgamma ligands suppresses growth of subgroup of these cell lines. However, expression and subcellular distribution of PPARgamma in human gastric cancer tissues is still unknown. Therefore, expression and subcellular localization of PPARgamma were examined among different histological types of gastric cancer tissues. Immunohistochemical staining for PPARgamma was performed using biopsy specimens of human gastric cancer of various histological types, gastric adenomas, and intestinal metaplasia. All samples of intestinal metaplasia and most samples of gastric tumors, except for signet ring cell carcinoma, expressed PPARgamma in the epithelial cells. Most samples of signet ring cell cancer lacked PPARgamma expression. All samples of intestinal metaplasia expressed PPARgamma only in the cytosol. For adenoma, 90% was positive for PPARgamma in cytosol, and 40% was positive in nuclei, for well-differentiated adenocarcinoma, 80% was positive in cytosol, and 20% was positive in nuclei. For moderately differentiated adenocarcinomas, 70% was positive for cytosol, and 80% was positive for nuclei; for poorly differentiated adenocarcinoma, 30% was positive in cytosol, and 70% was positive in nuclei. The frequency of samples with positive cytosolic staining decreased as the differentiation stage turned from intestinal metaplasia to adenoma, well-, moderately-, and poorly-differentiated cancers. Simultaneously, there was a tendency toward an increased frequency of samples with positive nuclear PPARgamma staining as the differentiation stage transformed from intestinal metaplasia to poorly-differentiated cancer. There was a striking difference in subcellular localization according to the differentiation levels of gastric dysplastic cells. The findings also supported an intestinal metaplasia-adenoma-well-differentiated gastric cancer sequence, and signet ring cell cancer was suggested to be of a different lineage from other types of gastric cancers.

Ex-vivo study of high-magnification chromoendoscopy in the gastrointestinal tract to determine the optimal staining conditions for endocytoscopy.

BACKGROUND AND STUDY AIMS: Endocytoscopy allows the observation of living cells in the gastrointestinal tract. Consistently clear views are essential for clinical application of the technique, but these are not always obtained. The aim of this study was to determine an appropriate staining regimen for endocytoscopy. MATERIALS AND METHODS: This was an ex-vivo animal study in which we stained freshly resected porcine esophagus, stomach, and colon with different concentrations of three dyes (1%, 0.5%, and 0.25% crystal violet; 5%, 2.5%, and 1% methylene blue; and 1%, 0.5%, and 0.25% toluidine blue) and assessed them after different exposure times (10 seconds, 30 seconds, 60 seconds, and 90 seconds). The images obtained were evaluated according to the staining status of the cytoplasm and the nucleus, and the contrast between the cytoplasm and the nuclei, and the optimal staining conditions for each organ were determined. Additionally, freshly resected human esophagus, stomach, and colon tissues were tested under the dye/exposure conditions that were found to be the most appropriate in the animal study. RESULTS: After intensive mucus removal, high-quality images were obtained using methylene blue and toluidine blue. The optimum conditions for endocytoscopic observation were obtained after staining with 1% methylene blue in the esophagus and with 0.25% toluidine blue in the stomach and the colon, after 60 seconds of exposure to the dye. This was confirmed in the human specimens. CONCLUSIONS: This study provides important information on appropriate staining conditions for endocytoscopy. Further ex-vivo and in-vivo studies are necessary before this technique comes into standard use, however.

Manipulation of the small intestine as a cause of the increased inflammatory response after open compared with laparoscopic surgery.

BACKGROUND: Laparoscopic surgery of the gastrointestinal tract involves a reduced immune response compared with open surgery. The aim of this study was to assess manual handling of the gut in open procedures as the principal cause of the enhanced immune response. METHODS: Eighteen Landrace pigs underwent gastrectomy by three different methods: conventional open wound with bowel manipulation, laparoscopically assisted gastrectomy, and gastrectomy without manipulation using a combination of open wound and laparoscopic surgical devices. Local inflammatory changes were assessed by ascites formation, intestinal adhesion development and intestinal inflammatory gene expression. Associated systemic inflammatory changes were determined by measuring portal and systemic plasma endotoxin levels, plasma inflammatory cytokine levels, liver inflammatory gene expression and transaminase levels. RESULTS: Significantly more postoperative intra-abdominal fluid and adhesions were seen in the open group. The expression of inflammatory cytokines was significantly greater in the intestine and liver in the open group. Portal and systemic levels of endotoxin, inflammatory cytokines and transaminases were also higher. CONCLUSION: Manual handling of organs during gastrectomy is an important contributor to the molecular and humoral inflammatory response to surgery, supporting the use of minimally invasive techniques in gastrointestinal surgery.

Strong association of ARK5 with tumor invasion and metastasis.

We recently identified a novel human AMPK family member, ARK5, and discovered that is a major factor in Akt-dependent cancer cell survival and migration activity through activation of MT1-MMPs in vitro. The mRNA expression of other AMPK family members and ARK5 was measured using RT-PCR in human colorectal carcinoma cell lines DLD-1, WiDr, HCT-15, SW620, LoVo, SW480, and mRNA expression of AMPK-alpha1, SNARK, MELK and ARK5, but not AMPK-alpha2, was detected in every line. Quantitative-PCR (Q-PCR) to estimate the amount of ARK5 mRNA expression in the cell lines showed that there is a variety of ARK5 expressions among the cell lines and high expression was observed in a cell line derived from the metastatic lesion, LoVo. To determine the effect of ARK5 overexpression on metastasis in vivo, we established human pancreas cancer cell line PANC-1 stably transfected with ARK5 full-length expression vector (P/ARK) and DLD-1 stably transfected with the same vector (D/ARK). Migration assay showed a remarkable increase in the activity both in P/ARK and D/ARK, and an in vivo metastasis assay showed a marked increase of P/ARK in liver metastasis. Based on these observations, it is suggested that ARK5 expression is involved in cancer invasion and metastasis.

A seasonal variation in the onset of postoperative adhesive small bowel obstruction is related to changes in the climate.

BACKGROUND: Postoperative small bowel obstruction following abdominal procedures is more common in patients who have undergone laparotomy. However, little is known about the influence of climate on the incidence of postoperative small bowel obstruction. METHODS: To evaluate whether seasonal climatic variations are a risk factor for postoperative small bowel obstruction, hospital-based, retrospective case series was designed from medical records of 230 patients suffering from postoperative small bowel obstruction admitted to the Tokyo University Branch Hospital. Detailed analysis of weather charts from the Japanese Meteorological Agency and review of medical records for selected patients who were diagnosed with postoperative small bowel obstruction. The obstruction was diagnosed by abdominal X-ray imaging, clinical examination, and patient interviews. RESULTS: A total of 233 patients diagnosed with postoperative small bowel obstruction were identified. Analysis of the medical records of these 233 patients revealed that the variables associated with an increased risk of postoperative small bowel obstruction included low ambient temperatures of 5-10 degrees C, an increase in air humidity by 40-50% and air pressure of 1010-1015 hPa. CONCLUSION: The typical winter weather in Tokyo is characterised by low temperatures, low humidity and moderate air pressure. These winter climate conditions could be correlated with an increased incidence of postoperative small bowel obstruction in Tokyo during our period.

Low concentrations of ethanol induce apoptosis in human intestinal cells.

BACKGROUND: Increased systemic levels of endotoxin have been detected in human alcoholics and are thought to be derived from the gut. Although a 'leaky gut' is considered to be a necessary factor for alcohol-induced endotoxemia followed by chronic liver injury, the effects of low concentrations of ethanol on intestinal epithelial cells have not been fully understood. The aim of this study was to evaluate intestinal epithelial cell death induced by acute, low concentrations of ethanol in an in vitro system. METHODS: The human intestinal Caco-2 cell line was incubated with 0%, 5%, 10% ethanol for up to 3 h. Phosphatidylserine (PS) externalization, caspase-mediated cytokeratin 18 (CK18) cleavage, and DNA fragmentation were evaluated using flow cytometry. The caspase inhibitor zVAD-fmk was used to test the role of caspases in ethanol-induced cell death. RESULTS: Treatment with 5% and 10% ethanol for 3 h led to a gradual increase in PS externalization. Caspase-mediated CK18 was significantly enhanced as early as 1 h after 10% ethanol incubation, while DNA fragmentation was detected from 2 h onwards. Not only caspase activation but also both PS externalization and DNA fragmentation were completely prevented by pretreatment with the caspase inhibitor. CONCLUSIONS: Apoptotic cell death in confluent Caco-2 cells was induced by acute and low concentrations of ethanol. These results suggest that clinically achievable doses of ethanol impair intestinal barrier function by induction of apoptosis in intestinal epithelial cells. This impairment of the barrier function would allow endotoxin to enter the circulation and evoke hepatic inflammation.

Renal denervation abolishes the protective effects of ischaemic preconditioning on function and haemodynamics in ischaemia-reperfused rat kidneys.

Studies were conducted to investigate the role of renal sympathetic nerves in the process of acquiring ischaemic tolerance in ischaemic preconditioned ischaemia-reperfused rat kidneys. Two periods of 3-min occlusion of bilateral renal arteries was performed prior to 30-min bilateral ischaemia and 90-min reperfusion in acute renal denervated or innervated kidneys. The glomerular filtration rate (GFR), fractional excretion of sodium (FENa) and lithium (FELi), and renal blood flow (RBF) were assessed in reperfused kidneys. Ischaemic preconditioning significantly improved values for all these parameters as compared with no treated ischaemia-reperfused kidneys. Denervation caused slight increase in GFR, diuresis and natriuresis without improving RBF after reperfusion. However, protecting effects of ischaemic preconditioning on renal function were disappeared in denervated kidneys, while in innervated kidneys the effects of ischaemic preconditioning were maintained. These results clearly showed that ischaemic preconditioning pre-treatment protects kidneys against ischaemia-reperfusion injury, and the effects are, at least in part, mediated by sympathetic nerves, as the protective effects were abolished by denervation.

Immunohistochemical prognostic indicators of gastrointestinal carcinoid tumours.

AIMS: The aim of this study was to determine whether expression of the oncoproteins p21, p53, E-cadherin (EC), cyclin D1, bcl-2 and Rb and the proliferation marker Ki-67 is predictive of malignant behaviour in gastrointestinal carcinoid tumours. METHODS: Immunohistochemical (IHC) staining was performed on carcinoid tumours from 41 patients (31 rectal, eight gastrointestinal, two appendiceal lesions). The six tumours that had invaded deeply into the muscularis propria or beyond, had metastasized to regional lymph nodes or had metastasized to a distant site were classified as the malignant group, and the other 35 tumours formed the benign group. IHC expression was compared between the two groups, and the prognostic value of each marker was assessed. RESULTS: Of the six tumours in the malignant group, 66.7% were p21 positive, 0% were p53 positive, 33.3% were EC positive, 100% were cyclin D1 positive, 33.3% were Rb positive, 16.7% were bcl-2 positive and 50% were Ki-67 positive. Of the 35 tumours in the benign group, 17.1% were p21 positive, 0% were p53 positive, 100% were EC positive, 94.3% were cyclin D1 positive, 8.6% were Rb positive, 17.1% were bcl-2 positive and 0% were Ki-67 positive. CONCLUSIONS: These data show that p53, cyclin D1, Rb, bcl-2 and Ki-67 staining does not correlate with malignant behaviour but that overexpression of p21 (P=0.02) and reduced staining of EC (P=0.005) do correlate with malignant behaviour. These two parameters may therefore be useful as prognostic indicators for gastrointestinal carcinoid tumours.

Pathophysiological relevance of the CD14 receptor in surgical patients: biological activity of endotoxin is regulated by the CD14 receptor.

Endotoxins (lipopolysaccharides, LPSs) are potent bacterial poisons, and they are always present in the intestine in considerable numbers. Stress, such that as a resulting from multiple injuries, burns, hypovolemia, hypoxia, intestinal ischemia, and surgery can lead to a breakdown of the gut barrier, allowing endotoxins to enter the systemic circulation via translocation. However, estimating the biological activity of translocated circulating endotoxins and identification of the mechanisms regulating their biological activities remain complex problems. CD14 has been found to exist as a soluble protein in the serum and as a glycosylphosphatidylinositol (GPI)-anchored protein of myeloid lineage cells. It plays key roles in both LPS-induced activation and in LPS internalization by cells. In this article, we outline: (i) the biological activity of circulating endotoxin; and (ii) the role of membrane and/or soluble CD14 regulating the bioactivity of circulating endotoxin in a human model of postoperative endotoxemia.

Lymph node metastasis and preoperative diagnosis of depth of invasion in early gastric cancer.

BACKGROUND: No reports have, to date, focused on the relationship between preoperative determination of the depth of invasion and lymph node metastasis. The present study, under the leadership of the Japanese Gastric Cancer Association, was designed to form a basis for decision making in limited treatment for early gastric cancer (EGC). METHODS: From eight major hospitals in Japan, 2672 gastric cancers whose preoperative depth of invasion was mucosal(M-cancer), and 6209 EGCs, consisting of 3584 mucosal(m-) and 2625 submucosal(sm-) cancers, were collected by questionnaire. All registered patients underwent gastrectomy with D1 or more extensive lymphadenectomy between 1985 and 1998. RESULTS: The accuracy of preoperative diagnosis of depth of invasion of M-cancers was 80.2% (2144/2672). However, of the total of 2432 M-cancers in which no nodal involvement was observed intraoperatively (N0), histological examination of the resected specimens confirmed that lymph node metastasis was absent in 2353 (96.8%). The frequencies of lymph node metastasis in early gastric, m-, and sm-cancers were 8.9%, 2.5%, and 17.6%, respectively. Node involvement was associated with a higher frequency of undifferentiated than differentiated histology, as well as with greater tumor size. The incidences of lymph node metastasis in m-cancers with a diameter of less than 4 cm, and in sm-cancers with a diameter below 1 cm were 1.3% (37/2837) and 4.9% (4/82), respectively. These metastases rarely extended beyond the first tier. CONCLUSION: N0 and M-cancers, m-cancers less than 4 cm in diameter, and sm-cancers no larger than 1 cm in diameter may be appropriate indications for limited surgery.

Gender differences in cytokine secretion by human peripheral blood mononuclear cells: role of estrogen in modulating LPS-induced cytokine secretion in an ex vivo septic model.

Clinical studies demonstrate a better outcome of sepsis in females. Elevated estrogen levels and plasma cytokine imbalance occur in septic patients. We propose that gender-different cytokine secretion by the peripheral blood mononuclear cells (PBMCs) in sepsis determines the clinical outcome. A 2 x 10(6) PBMC sample from healthy volunteers (10 males and 10 females) was incubated with 1 ng/mL of lipopolysaccharide (LPS), estradiol (E2; 0, 0.03, 0.3, 3.0, 30 ng/mL), or 1 ng/mL of LPS + E2 (0, 0.03, 0.3, 3.0, 30 ng/ml), and supernatant cytokine levels were measured. Tumor necrosis factor alpha (TNF alpha) and interleukin (IL)-6 production by PBMCs from both sexes was time-dependently stimulated by LPS. At 6 h after LPS challenge, the TNF alpha level of male PBMCs was significantly higher but IL-6 secretion by female PBMCs was higher (two-way ANOVA: P < 0.05). E2 alone stimulated cytokine secretion by male PBMCs. Addition of the same E2 concentration as in sepsis patients' plasma modulated LPS-induced cytokine production. No significant sex differences in LPS-stimulated TNF alpha or IL-6 secretion by PBMCs were found, but IL-10 secretion by male PBMCs was significantly suppressed. This study demonstrated a gender difference in PBMCs responsiveness to LPS and E2 stimulation and E2-modulated cytokine secretion. In this PBMCs model of sepsis, only the supernatant IL-10 level was significantly lower in males. These ex vivo findings may partially explain the mechanism underlying the poorer outcome of male sepsis patients.