Preoperative chemoradiation therapy for esophageal cancer is a risk factor for the elevation of high mobility group box-1, leading to an increase in postoperative severe pulmonary complications.

We herein clarified the time course of changes in the serum high mobility group box chromosomal protein-1 (HMGB-1) concentrations in esophageal cancer patients after esophagectomy, and investigated whether the perioperative serum HMGB-1 levels correlate with the administration of neoadjuvant chemoradiation therapy (NACRT) and the postoperative clinical course, especially the occurrence of pulmonary complications, in such patients. Sixty patients who underwent right transthoracic esophagectomy for esophageal cancer were enrolled in this study. The relationship between the perioperative serum HMGB-1 levels and NACRT, and the postoperative severe pulmonary complications were evaluated. Patients with severe pulmonary complications (n = 44) tended to have undergone NACRT more often than those without severe pulmonary complications (n = 16). The preoperative and postoperative day 7 serum HMGB-1 concentrations were significantly higher in patients with severe pulmonary complications than those in patients without severe pulmonary complications. In the univariate and multivariate analyses, the use of NACRT and the preoperative elevations in the serum HMGB-1 levels (>4.2 ng/mL) were found to be significantly associated with pulmonary dysfunction. Furthermore, the response to NACRT was found to be significantly associated with the preoperative serum HMGB-1 levels. The use of NACRT contributes to preoperative serum HMGB-1 elevation, and these were risk factors for the occurrence of severe postoperative pulmonary complications in patients with esophageal cancer after thoracic esophagectomy.

The AURKA/TPX2 axis drives colon tumorigenesis cooperatively with MYC.

BACKGROUND: The MYC oncogene has long been established as a central driver in many types of human cancers including colorectal cancer. However, the realization of MYC-targeting therapies remains elusive; as a result, synthetic lethal therapeutic approaches are alternatively being explored. A synthetic lethal therapeutic approach aims to kill MYC-driven tumors by targeting a certain co-regulator on the MYC pathway. PATIENTS AND METHODS: We analyzed copy number and expression profiles from 130 colorectal cancer tumors together with publicly available datasets to identify co-regulators on the MYC pathway. Candidates were functionally tested by in vitro assays using colorectal cancer and normal fibroblast cell lines. Additionally, survival analyses were carried out on another 159 colorectal cancer patients and public datasets. RESULTS: Our in silico screening identified two MYC co-regulator candidates, AURKA and TPX2, which are interacting mitotic regulators located on chromosome 20q. We found the two candidates showed frequent co-amplification with the MYC locus while expression levels of MYC and the two genes were positively correlated with those of MYC downstream target genes across multiple cancer types. In vitro, the aberrant expression of MYC, AURKA and TPX2 resulted in more aggressive anchorage-independent growth in normal fibroblast cells. Furthermore, knockdown of AURKA or TPX2, or treatment with an AURKA-specific inhibitor effectively suppressed the proliferation of MYC-expressing colorectal cancer cells. Additionally, combined high expression of MYC, AURKA and TPX2 proved to be a poor prognostic indicator of colorectal cancer patient survival. CONCLUSIONS: Through bioinformatic analyses and experiments, we proposed TPX2 and AURKA as novel co-regulators on the MYC pathway. Inhibiting the AURKA/TPX2 axis would be a novel synthetic lethal therapeutic approach for MYC-driven cancers.

Prognostic impact of histological categorisation of epithelial-mesenchymal transition in colorectal cancer.

BACKGROUND: The crosstalk between cancer cells and stroma is involved in the acquired capability for metastasis through the induction of epithelial-mesenchymal transition (EMT). We aimed to clarify the prognostic value of the histological category of EMT in colorectal cancer (CRC). METHODS: Tumour EMT was graded into one of three histological categories on the basis of integrated assessment of poorly differentiated clusters and pro-EMT desmoplasia at the leading edge of the primary tumour (Histology(EMT)). Stage II and III CRC patients (cohort 1, N=500) and stage IV patients (cohort 2, N=196) were retrospectively analysed. RESULTS: In cohort 1, patients were stratified into three groups with widely different disease-free survival rates (95%, 83% and 39%) on the basis of Histology(EMT) (P<0.0001). In cohort 2, Histology(EMT) significantly stratified overall survival of patients irrespective of metasectomy. Multivariate analyses indicated that Histology(EMT) had a strong prognostic impact independent of staging factors. Statistically, Histology(EMT) had a better prognostic stratification power than T and N stages; however, in cohort 2, the power of M substage was superior. CONCLUSIONS: A histological model to categorise EMT by integrated assessment of dedifferentiation and desmoplastic environment is a potent prognostic index independent of staging factors.

Protective effect of the tunneling nanotube-TNFAIP2/M-sec system on podocyte autophagy in diabetic nephropathy.

Podocyte injury leading to albuminuria is a characteristic feature of diabetic nephropathy (DN). Hyperglycemia and advanced glycation end products (AGEs) are major determinants of DN. However, the underlying mechanisms of podocyte injury remain poorly understood. The cytosolic protein TNFAIP2/M-Sec is required for tunneling nanotubes (TNTs) formation, which are membrane channels that transiently connect cells, allowing organelle transfer. Podocytes express TNFAIP2 and form TNTs, but the potential relevance of the TNFAIP2-TNT system in DN is unknown. We studied TNFAIP2 expression in both human and experimental DN and the renal effect of tnfaip2 deletion in streptozotocin-induced DN. Moreover, we explored the role of the TNFAIP2-TNT system in podocytes exposed to diabetes-related insults. TNFAIP2 was overexpressed by podocytes in both human and experimental DN and exposre of podocytes to high glucose and AGEs induced the TNFAIP2-TNT system. In diabetic mice, tnfaip2 deletion exacerbated albuminuria, renal function loss, podocyte injury, and mesangial expansion. Moreover, blockade of the autophagic flux due to lysosomal dysfunction was observed in diabetes-injured podocytes both in vitro and in vivo and exacerbated by tnfaip2 deletion. TNTs allowed autophagosome and lysosome exchange between podocytes, thereby ameliorating AGE-induced lysosomal dysfunction and apoptosis. This protective effect was abolished by tnfaip2 deletion, TNT inhibition, and donor cell lysosome damage. By contrast, Tnfaip2 overexpression enhanced TNT-mediated transfer and prevented AGE-induced autophagy and lysosome dysfunction and apoptosis. In conclusion, TNFAIP2 plays an important protective role in podocytes in the context of DN by allowing TNT-mediated autophagosome and lysosome exchange and may represent a novel druggable target.Abbreviations: AGEs: advanced glycation end products; AKT1: AKT serine/threonine kinase 1; AO: acridine orange; ALs: autolysosomes; APs: autophagosomes; BM: bone marrow; BSA: bovine serum albumin; CTSD: cathepsin D; DIC: differential interference contrast; DN: diabetic nephropathy; FSGS: focal segmental glomerulosclerosis; HG: high glucose; KO: knockout; LAMP1: lysosomal-associated membrane protein 1; LMP: lysosomal membrane permeabilization; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; PI3K: phosphoinositide 3-kinase; STZ: streptozotocin; TNF: tumor necrosis factor; TNFAIP2: tumor necrosis factor, alpha-induced protein 2; TNTs: tunneling nanotubes; WT: wild type.

Evaluation of the seventh edition of the tumour, node, metastasis (TNM) classification for colon cancer in two nationwide registries of the United States and Japan.

AIM: The new TNM classification is currently being implemented. We evaluated the TNM-7 staging system based on the two nationwide colon cancer registries in the United States and Japan to clarify whether this system better stratifies patients' prognoses than the TNM-6 did and to determine whether stratification can be effectively simplified. METHODS: The Surveillance, Epidemiology, and End Results population-based data from 1988 to 2001 for 50139 colon cancer patients and the multi-institutional registry data from the Japanese Society for Cancer of the Colon and Rectum from 1984 to 1994 for 10754 patients were analysed. We devised a modified version of the TNM-7 staging system to allow simpler classification of the TN categories and compared the TNM-6, TNM-7, modified TNM-7, and the Dukes staging system based on survival curves and objective statistical tests such as likelihood ratio χ(2) tests, Akaike's information criterion, and Harrell's c-index. RESULTS: The TNM-7 was superior to the TNM-6 in all objective statistical tests in the United States (c-index; 0.700 vs 0.696, P<0.001) as well as in the Japan data sets (0.732 vs 0.729, P=0.035). The modified TNM-7 is much simpler, but it nevertheless showed similar values to those of the original TNM-7 (c-index; the United States 0.702, Japan 0.733). CONCLUSIONS: The new TNM-7 is complicated but better at stratifying patients than the TNM-6 in the United States and Japan, and could be effectively simplified.

Optimal margins and lymphadenectomy in colonic cancer surgery.

BACKGROUND: A standard management policy has not yet been established with respect to the extent of lymphadenectomy for colonic cancer. METHODS: A total of 914 consecutive patients who underwent potentially curative surgery for T2-T4 colonic cancer were reviewed retrospectively. The number of lymph nodes (LNs) examined and the potential contributions to the staging accuracy of the distinct area were analysed. The survival benefit of dissection was compared for pericolic (local), mesocolic (intermediate) and main arterial trunk (main) LN. RESULTS: Removal of the pericolic LNs within 5 cm of the tumour and intermediate LNs resulted in a mean LN number of 15.9, a sensitivity for overall node positivity of 97.5 per cent, and a survival benefit calculated as a therapeutic value index of 31.4 points. The additional removal of LNs more than 5 cm from the tumour and main LNs did not improve the staging accuracy, while adding only 3.4 points to the survival benefit. CONCLUSION: Current guidelines may encourage needlessly extensive surgery. Clinical trials to establish the optimal extent of lymphadenectomy are warranted.

Prognostic and predictive values of tumour budding in stage IV colorectal cancer.

BACKGROUND: Tumour budding is an important prognostic feature in early-stage colorectal cancer, but its prognostic significance in metastatic disease has not been fully investigated. METHODS: Patients with stage IV disease who had primary colorectal tumour resection without previous chemotherapy or radiotherapy from January 2000 to December 2018 were reviewed retrospectively. Budding was evaluated at the primary site and graded according to the criteria of the International Tumor Budding Consensus Conference (ITBCC) (BD1, low; BD2, intermediate; BD3, high). Patients were categorized by metastatic (M1a, M1b) and resectional (R0/R1, R2/unresected) status. Subgroups were compared for overall (OS) and recurrence-free (RFS) survival in R0/R1 subgroups; R2/unresected patients were evaluated for the rate of tumour progression, based on change in tumour size from baseline. RESULTS: Of 371 patients observed during the study, 362 were analysed. Patients with BD3 had a lower 5-year OS rate than those with BD1 + BD2 (18·4 versus 40·5 per cent; P < 0·001). Survival analyses according to metastatic and resection status also showed that BD3 was associated with shorter OS than BD1 + BD2. In multivariable analysis, BD3 (hazard ratio (HR) 1·51, 95 per cent c.i. 1·11 to 2·10; P = 0·009), T4 status (HR 1·39) and R2/unresected status (HR 3·50) were associated with decreased OS. In the R0/R1 subgroup, the 2-year RFS rate was similar for BD3 and BD1 + BD2 according to metastatic status. There was no significant difference between BD3 and BD1 + BD2 for change in tumour size in the R2/unresected subgroup (P = 0·094). Of 141 patients with initially unresectable metastases who had chemotherapy, 35 achieved conversion from unresectable to resectable status. The conversion rate was significantly higher for BD1 + BD2 than for BD3 (36 versus 18 per cent; P = 0·016). CONCLUSION: Stage IV colorectal cancer with high-grade tumour budding according to ITBCC criteria correlates with poor prognosis.

ANTECEDENTES: La esofaguectomía por cáncer se asocia con un descenso de la calidad de vida relacionada con la salud (health-related quality of life, HRQoL) a largo plazo. El objetivo de este estudio fue evaluar el efecto de las comorbilidades sobre la HRQOL entre pacientes supervivientes de cánceres de esófago o de la unión gastroesofágicas después de 10 años o más. MÉTODOS: Este estudio incluye una cohorte de base poblacional recogida de forma prospectiva que incluía todos los pacientes operados de cáncer de esófago o de la unión gastroesofágica en Suecia en 2001-2005 con seguimiento hasta el 31 de diciembre de 2016. Todos los datos relacionados con las características de los pacientes y del tumor, detalles del tratamiento y HRQoL se recogieron en una base de datos prospectiva. Se utilizaron modelos de regresión multivariable ANCOVA, ajustados por edad, sexo, histología del tumor, estadio, y técnica quirúrgica, para calcular las puntuaciones medias ajustadas con los i.c. del 95% para todas las variables de la HRQoL. RESULTADOS: Un total de 92 (88%) supervivientes respondieron a los cuestionarios. En función del impacto de las comorbilidades en la salud en general, se clasificaron a los pacientes en los grupos de bajo versus alto impacto. Los resultados muestran que los pacientes en el grupo de alto impacto presentaban un descenso clínicamente significativo de la HRQoL y un aumento en el nivel de síntomas, pero las diferencias entre estos dos grupos no fueron estadísticamente significativas. CONCLUSIÓN: A los 10 años de la esofaguectomía por cáncer, las comorbilidades con un alto impacto sobre la salud general siguen contribuyendo en el deterioro de la HRQoL.

Predictive immunohistochemical features for tumour response to chemoradiotherapy in rectal cancer.

BACKGROUND: Reduced expression of cluster of differentiation (CD) 133 and cyclo-oxygenase (COX) 2, and increased density of CD8+ tumour-infiltrating lymphocytes, are associated with a favourable tumour response to preoperative chemoradiotherapy (CRT). This study aimed to evaluate these markers in relation to tumour response after preoperative CRT in two rectal cancer cohorts. METHODS: Patients with low rectal cancer who underwent radical resection and preoperative short-term CRT in 2001-2007 (retrospective cohort) and long-term CRT in 2011-2017 (prospective cohort) were analysed. Pretreatment biopsies were stained immunohistochemically using antibodies to determine CD133 and COX-2 expression, and increased CD8+ density. Outcome measures were tumour regression grade (TRG), tumour downstaging and survival. RESULTS: For 95 patients in the retrospective cohort, the incidence of TRG 3-4 was 67 per cent when two or three immunohistochemistry (IHC) features were present, but only 20 per cent when there were fewer features (P < 0·001). The incidence of tumour downstaging was higher in patients with at least two IHC features (43 versus 22 per cent with fewer features; P = 0·029). The 49 patients in the prospective cohort had similar rates to those in the retrospective cohort (TRG 3-4: 76 per cent for two or more IHC features versus 25 per cent with fewer features, P < 0·001; tumour downstaging: 57 versus 25 per cent respectively, P = 0·022). Local recurrence-free survival rates in patients with more or fewer IHC features were similar in the retrospective and prospective cohort (P = 0·058 and P = 0·387 respectively). CONCLUSION: Assessment of CD133, COX-2 and CD8 could be useful in predicting a good response to preoperative CRT in patients with lower rectal cancer undergoing neoadjuvant therapy. Further studies are needed to validate the results in larger cohorts and investigate a survival benefit.

ANTECEDENTES: La expresión reducida de CD133 and COX-2, y un aumento en la densidad de los linfocitos infiltrantes del tumor CD8+ se han asociado recientemente con una respuesta favorable del tumor a la quimiorradioterapia preoperatoria (preoperative chemoradiotherapy, CRT). Este estudio evaluó estos marcadores respecto a la respuesta del tumor tras CRT preoperatoria en dos cohortes de cáncer colorrectal. MÉTODOS: Se analizaron pacientes con cáncer de recto bajo sometidos a resección radical y CRT preoperatoria de corta duración entre 2001-2007 (cohorte retrospectiva) y CRT de larga duración entre 2011-2017 (cohorte prospectiva). Se realizó tinción inmunohistoquímica (immunohistochemical, IHC) con anticuerpos para CD133, COX-2 y CD8 en las biopsias previas al tratamiento. Las características de interés incluyeron la disminución en las expresiones de CD133 y COX-2, y la densidad aumentada de CD8+. Las variables de interés fueron los grados de regresión tumoral (tumour regression grades, TRG) de acuerdo con Rödel, la reducción del estadio tumoral y las supervivencias. RESULTADOS: La cohorte retrospectiva incluyó 95 pacientes. En este subgrupo, la incidencia de TRGs 3-4 fue del 66,7% en pacientes con dos o tres características de la IHC, mientras que solo fue del 20,0% en pacientes con ninguna o con una característica (P < 0,001). Además, la incidencia de disminución del estadio tumoral fue más alta en pacientes que mostraban al menos dos características IHC (43,3%) que en los controles (21,5%; P = 0,029). En la cohorte prospectiva se incluyeron 49 pacientes y la incidencia de estos hallazgos fue similar (TRG 3-4, 76,2% en ≥ 2 características IHC versus 25,0% en los controles, P < 0,001; disminución del estadio tumoral, 57,1% en ≥ 2 características IHC versus 25,0% en los controles, P = 0,022). La supervivencia libre de recidiva local fue similar en las cohortes retrospectiva y prospectiva, cuando se compararon subgrupos de acuerdo con las características IHC (P = 0,058 y 0,387, respectivamente) CONCLUSIÓN: Este estudio sugiere que la evaluación de CD133, COX-2 y CD8 podría ser útil para la predicción de una buena respuesta a la CRT preoperatoria en pacientes con cáncer de recto bajo sometidos a tratamiento neoadyuvante. Se necesitan estudios adicionales para validar los resultados en amplias cohortes e investigar el beneficio en la supervivencia.

Growth pattern in the muscular layer reflects the biological behaviour of colorectal cancer.

OBJECTIVE: To determine the clinical value of evaluating the cancer morphology in muscularis propria (MP) for colorectal cancer (CRC) patients. METHOD: A total of 994 patients with advanced CRC were reviewed in terms of two distinctive growth patterns in the MP: (i) horizontal spread between the circular and longitudinal muscle layers (H-spread) and (ii) 'streaming' spread between the muscle bundles of the circular muscle layer (S-spread). RESULTS: The incidence of H-spread (n = 153) and S-spread (n = 150) showed a positive correlation with tumour-node-metastasis (TNM) stage and both exerted a negative impact on postoperative survival. Adverse morphology in the MP (H-spread and/or S-spread) was consistent with a high grade of vascular invasion and budding in the extramural layer, as also with unfavourable fibrotic stromas in the reactive fibrous zone; the 5-year survival rate in patients with such features was 64.2%, which was lower than that in those without (86.5%, P < 0.0001). Multivariate analysis demonstrated that adverse morphology was an independent prognostic determinant, along with T- and N -stage. As the mode of H-spread, perineural invasion in the myenteric plexus was found to be predominant over lymphatic spread on the basis of S100 and CD34 immunostaining, but neural cell adhesion molecule expression, whether on cancer cells or on neural cells, was not significant for this growth pattern. CONCLUSION: A particular group of CRCs ingeniously utilizes the thin space between muscle fascicles for development in the MP. Although the biological mechanism remains unknown, this distinctive growth pattern could be a useful indicator to identify CRC patients at high risk of recurrence.

Gut commensal Bacteroides acidifaciens prevents obesity and improves insulin sensitivity in mice.

In humans, the composition of gut commensal bacteria is closely correlated with obesity. The bacteria modulate metabolites and influence host immunity. In this study, we attempted to determine whether there is a direct correlation between specific commensal bacteria and host metabolism. As mice aged, we found significantly reduced body weight and fat mass in Atg7ΔCD11c mice when compared with Atg7f/f mice. When mice shared commensal bacteria by co-housing or feces transfer experiments, body weight and fat mass were similar in both mouse groups. By pyrosequencing analysis, Bacteroides acidifaciens (BA) was significantly increased in feces of Atg7ΔCD11c mice compared with those of control Atg7f/f mice. Wild-type C57BL/6 (B6) mice fed with BA were significantly more likely to gain less weight and fat mass than mice fed with PBS. Of note, the expression level of peroxisome proliferator-activated receptor alpha (PPARα) was consistently increased in the adipose tissues of Atg7ΔCD11c mice, B6 mice transferred with fecal microbiota of Atg7ΔCD11c mice, and BA-fed B6 mice. Furthermore, B6 mice fed with BA showed elevated insulin levels in serum, accompanied by increased serum glucagon-like peptide-1 and decreased intestinal dipeptidyl peptidase-4. These finding suggest that BA may have potential for treatment of metabolic diseases such as diabetes and obesity.

Epidermal growth factor receptors on cultured neoplastic human thyroid cells and effects of epidermal growth factor and thyroid-stimulating hormone on their growth.

Epidermal growth factor (EGF) receptors on primary-cultured human thyroid cells from 27 neoplasias (nine adenomas and 18 differentiated carcinomas) were analyzed and compared with those on the cultured nonneoplastic part of human thyroid cells. Total binding of 125I-EGF to the nonneoplastic part, adenoma, and carcinoma cells did not differ significantly. Scatchard analysis showed that the neoplastic human thyroid cells, like their adjacent nonneoplastic counterparts, consistently possessed EGF receptors with two components. In a paired study of five patients, the association constant of the carcinoma cells' high-affinity component (Ka1) was found to be significantly lower than that of adjacent nonneoplastic thyroid cells (P less than 0.05). Furthermore, a study of the cells from 18 carcinomas revealed that overall their Ka1s (4.15 +/- 0.82 x 10(9) M-1, mean +/- SEM) were significantly lower than those of adenoma cells (10.34 +/- 1.51 x 10(9) M-1, n = 9) and of nonneoplastic cells adjacent to them (8.32 +/- 0.84 x 10(9) M-1, n = 23). The difference in Ka1s for adenoma and nonneoplastic thyroid cells was not statistically significant. The number of receptor sites (Cmax) per cell was not significantly different in any of the three. Incorporation of [3H]thymidine (dThd) increased significantly in all kinds of thyroid cells examined following the addition of 10 nM EGF, and the paired study showed that the size of this increase was not significantly different in neoplastic and adjacent nonneoplastic cells. The addition of 300 microunits/ml of thyroid-stimulating hormone caused a significant increase in dThd incorporation by adenoma cells but not by carcinoma or nonneoplastic cells. Furthermore, combined treatment with EGF and thyroid-stimulating hormone additively promoted adenoma cell growth only. A close inverse relationship was observed between Ka1 and the stimulatory effect of EGF on the dThd uptake in both nonneoplastic thyroid cells and adenoma cells. Carcinoma cells also showed similar profiles, but Ka1 relative to dThd increases were much smaller than the other two.

Direct interaction between an antigen-specific B cell clone and an MHC class II-reactive helper T cell clone.

TP67.14 established by somatic hybridization is a 2,4,6-trinitrobenzenesulfonic acid (trinitrophenyl, TNP)-specific B cell clone with a receptor molecule for TNP on the cell membrane, and MS202 is an interleukin-2 (IL-2)-dependent T helper (Th) cell clone reactive to auto-MHC class II antigens (IAk and IEk) as previously reported. In the present study it was shown that MS202 considerably induced the maturation of TP67.14 into anti-TNP plaque-forming cells (PFCs), and this response was markedly augmented by the addition of TNP-keyhole limpet hemocyanin (KLH). Recombinant cytokines and the culture supernatant of MS202 with TP67.14 did not affect the generation of anti-TNP antibodies by TP67.14. Also, neither anti-IL-4 nor anti-IL-5 monoclonal antibody (mAb) inhibited the maturation of TP67.14 mediated by MS202. The differentiative effect of MS202 on TP67.14 was completely lost when each cell was separately cultured using a semipermeable membrane. Monoclonal antibodies against LFA-1 beta molecules significantly blocked the development of anti-TNP PFCs induced by MS202, as well as anti-IAk and anti-IEk mAbs. Interestingly, the plasma membrane-enriched fraction (PM) derived from MS202 exhibited much more differentiative effects on TP67.14 treated with TNP-KLH than PM from other T cell lines and concanavalin A-induced T lymphoblasts. In addition, TNP-conjugated PM from MS202 by itself induced a great number of anti-TNP PFCs. The present findings indicate that MS202 is capable of inducing the maturation of TP67.14, which is considered to represent a population of B cells with antigen specificity in a late lineage of B cell maturation, through direct cell contact but not soluble factors. This suggests that B cells with antigen specificity, in the presence of antigen, can be induced to mature into antibody-secreting cells through direct contact with Th cells; in this process surface major histocompatibility complex class II and lymphocyte function-associated antigen 1 (LFA-1) molecules are directly involved and the cell membrane derived from Th cells provides a transductional signal for maturation of B cells with antigen specificity in the presence of antigen.

Increased CCR4 expression in active systemic lupus erythematosus.

CC chemokine receptor (CCR)4 is selectively expressed on Th2-type T cells and has been shown to be responsible for Th2-dominant immune responses. In this study, we analyzed the expression of CCR4 in active systemic lupus erythematosus (SLE) patients by FACS analysis using anti-human CCR4 monoclonal antibody and determined the clinical relevance in this disease. Higher expression of CCR4 was found on peripheral blood CD4+ T lymphocytes of active SLE patients than was found with healthy controls and inactive SLE patients. The CCR4 expression significantly correlated with the SLE disease activity index (SLEDAI) scores. The expression was dramatically decreased after the corticosteroid therapy in parallel with a serum level of double-stranded DNA antibody and SLEDAI scores. Moreover, we found that serum levels of IL-10 were increased in active SLE patients and significantly correlated with the CCR4 expression. This study suggests that Th2 immune response is predominant in the active state of SLE, and CCR4 may have relevance in regard to the disease course in SLE patients.

Visualization of the entire differentiation process of murine M cells: suppression of their maturation in cecal patches.

The microfold (M) cell residing in the follicle-associated epithelium is a specialized epithelial cell that initiates mucosal immune responses by sampling luminal antigens. The differentiation process of M cells remains unclear due to limitations of analytical methods. Here we found that M cells were classified into two functionally different subtypes based on the expression of Glycoprotein 2 (GP2) by newly developed image cytometric analysis. GP2-high M cells actively took up luminal microbeads, whereas GP2-negative or low cells scarcely ingested them, even though both subsets equally expressed the other M-cell signature genes, suggesting that GP2-high M cells represent functionally mature M cells. Further, the GP2-high mature M cells were abundant in Peyer's patch but sparse in the cecal patch: this was most likely due to a decrease in the nuclear translocation of RelB, a downstream transcription factor for the receptor activator of nuclear factor-κB signaling. Given that murine cecum contains a protrusion of beneficial commensals, the restriction of M-cell activity might contribute to preventing the onset of any excessive immune response to the commensals through decelerating the M-cell-dependent uptake of microorganisms.

Role of liver-type glucose transporter (GLUT2) in transport across the basolateral membrane in rat jejunum.

To obtain information on the regulation of glucose transport across the basolateral membrane (BLM) of intestinal epithelial cells, we measured the number of [3H]cytochalasin B binding sites and the level of liver-type glucose transporter (GLUT2) protein in the BLM in the jejunum of rats (i) with diabetes (ii) given a high-carbohydrate diet or (iii) with experimental hyperglycemia (12 h infusion of a high-glucose solution). A glucose uptake and the number of D-glucose inhibitable [3H]cytochalasin B binding sites in BLM vesicles were significantly increased in all three conditions. Western blot analysis showed that the amount of GLUT2 protein in BLM vesicles was increased in rats with diabetes and those given a high-carbohydrate diet, but not in those with experimental hyperglycemia. These results suggest that there is a mechanism for rapid regulation of glucose transport in the BLM that does not depend on change in the amount of GLUT2.

Inhibitory effect of tetrahydroswertianolin on tumor necrosis factor-alpha-dependent hepatic apoptosis in mice.

We investigated the effect of tetrahydroswertianolin (THS), a hepatoprotective agent from Swertia japonica, on tumor necrosis factor-alpha (TNF-alpha)-dependent hepatic apoptosis induced by D-galactosamine (D-GalN) (700 mg/kg, i.p.) and lipopolysaccharide (LPS) (10 microg/kg, i.p.) in mice. Apoptotic symptoms were observed at the initial stage of liver damage. By 5 hr after intoxication, hepatic DNA fragmentation had risen to 2123%, with the value in untreated mice set at 100%, without a significant elevation of serum alanine transaminase (ALT) activity. There was a parallel increase in hepatocytes undergoing chromatin condensation and apoptotic body formation. By 8 hr after intoxication, serum ALT activity had risen to 3707 U/L. Pretreatment with THS (50 mg/kg, p.o.) at 18 and 2 hr before intoxication significantly reduced DNA fragmentation to 821% of that in untreated mice and prevented the emergence of chromatin condensation and apoptotic body formation. A significant and dose-dependent reduction in serum ALT activity at 8 hr also was observed with THS pretreatment. These effects of THS were different from those observed from pretreatment with glycyrrhizin (GCR), which is a clinically used hepatoprotective agent with membrane-stabilizing activity. GCR pretreatment (100 mg/kg, p.o.) did not inhibit hepatic DNA fragmentation (1588% of untreated mice), although this compound significantly protected against serum ALT elevation (1463 U/L). These data suggest that an inhibitory effect on the progression of hepatic apoptosis prior to liver injury may be involved in the hepatoprotective mechanisms of THS, whereas it appears that GCR affects the processes after apoptosis. In a separate experiment, we found that the concentration of serum TNF-alpha rose to 2016 pg/mL at 1 hr after intoxication of mice with D-GalN and LPS, but this increase was suppressed by THS pretreatment (10, 50, or 200 mg/kg, p.o.) to 716, 454, or 406 pg/mL, respectively. Further study with a reverse transcriptase-polymerase chain reaction method showed that THS blocked TNF-alpha production at the transcriptional level. Because TNF-alpha is a critical mediator to elicit apoptosis in this model, the property of suppressing TNF-alpha production may be of prime importance for THS inhibition of hepatic apoptosis.

Effect of simvastatin (MK-733) on plasma triacylglycerol levels in rats.

The effect of simvastatin (MK-733), an inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, on plasma triacylglycerol (TG) levels was studied in rats. Dietary administration of MK-733 (0.055%, w/w) for 7 days significantly (P less than 0.05) reduced plasma TG levels by 30.6% associated with a 44.3% significant (P less than 0.01) reduction in very low density lipoprotein TG (VLDL-TG) as compared to those in the concurrent control rats. Clofibrate (0.08%, w/w) also significantly (P less than 0.05) decreased plasma TG levels by 26.1%. MK-733 did not affect the triacylglycerol secretion rate (TGSR) during 0-1.5 hr after administration of Triton WR-1339, but reduced it by 33.9% during 1.5-3.0 hr. Clofibrate also decreased TGSR during 1.5-3.0 hr. MK-733 increased lipoprotein lipase (LPL) activity in epididymal adipose tissue and thigh muscle by 36.3 and 55.0% respectively. MK-733 significantly (P less than 0.05) increased LPL activity in the post-heparin plasma by 21.5%, although it did not affect hepatic triacylglycerol lipase (H-TGL) activity. Clofibrate did not affect LPL activity in the tissues or LPL and H-TGL activities in the post-heparin plasma. It is considered that the mechanism of plasma TG-lowering effect of MK-733 is the removal of VLDL-TG by an increase in LPL activity in the tissues as well as a decrease in the TGSR.

Inhibition of nitric oxide by phenylethanoids in activated macrophages.

Nitric oxide (NO) is one of the pro-inflammatory molecules. Some phenylethanoids have been previously shown to possess anti-inflammatory effects. Seven phenylethanoids from the stems of Cistanche deserticola, viz. isoacteoside, tubuloside B, acteoside, 2'-O-acetylacteoside, echinacoside, cistanoside A and tubuloside A, were tested for their effect on NO radical generation by activated murine macrophages. At the concentration of 100-200 microM, all the phenylethanoids reduced (6.3-62.3%) nitrite accumulation in lipopolysaccharide (0.1 microgram/ml)-stimulated J774.1 cells. At 200 microM, they inhibited by 32.2-72.4% nitrite accumulation induced by lipopolysaccharide (0.1 microgram/ml)/interferon-gamma (100 U/ml) in mouse peritoneal exudate macrophages. However, these compounds did not affect the expression of inducible nitric oxide (iNOS) mRNA, the iNOS protein level, or the iNOS activity in lipopolysaccharide-stimulated J774.1 cells. Instead, they showed a clear scavenging effect (6.9-43.9%) at the low concentrations of 2-10 microM of about 12 microM nitrite generated from an NO donor, 1-propanamine-3-hydroxy-2-nitroso-1-propylhydrazino (PAPA NONOate). These results indicate that the phenylethanoids have NO radical-scavenging activity, which possibly contributes to their anti-inflammatory effects.

Acteoside inhibits apoptosis in D-galactosamine and lipopolysaccharide-induced liver injury.

We assessed the effect of acteoside, a naturally occurring antioxidative phenylethanoid, on hepatic apoptosis and the subsequent liver failure induced by D-Galactosamine (D-GalN) and lipopolysaccharide (LPS). A co-administration of D-GalN (700 mg/kg) and LPS (35 microg/kg) to mice evoked typical hepatic apoptosis characterized by DNA fragmentation and apoptotic body formation, resulting in fulminant hepatitis and lethality of mice. Pre-administration of acteoside at 10 or 50 mg/kg subcutaneously at 12 and 1 h prior to D-GalN/LPS intoxication significantly inhibited hepatic apoptosis, hepatitis and lethality. Tumor necrosis factor-alpha (TNF-alpha) secreted from LPS-stimulated macrophages is an important mediator of apoptosis in this model. Acteoside showed no apparent effect on the marked elevation of serum TNF-alpha, but it partially prevented in vitro TNF-alpha (100 ng/ml)-induced cell death in D-GalN (0.5 mM)-sensitized hepatocytes at the concentrations of 50, 100 and 200 microM. These results indicated that D-GalN/LPS-induced hepatic apoptosis can be blocked by an exogenous antioxidant, suggesting the involvement of reactive oxygen intermediates (ROIs) in TNF-alpha-dependent hepatic apoptosis.

Assessment of clinicopathologic features of colorectal mucinous adenocarcinoma.

The purpose of this report is to analyze the clinicopathologic features of colorectal mucinous adenocarcinoma (MC), which is generally believed to have a poor prognosis, in an attempt to assess ways in which the surgical outcome can be improved. Clinicopathologic features of 44 patients with MC (6.6%), from among 662 patients with primary colorectal cancers, were compared with those of 545 patients with nonmucinous (non-MC) adenocarcinoma. MC is more likely to invade the adjacent viscera (29% versus 10%, p < 0.005) and show more extensive lymph node involvement beyond the pericolonic region (50% versus 26%, p < 0.005) than non-MC. Based on these findings, a more aggressive attitude toward surgical intervention is recommended, including extensive lymph node dissection and the resection of adjacent organs that seems to be affected macroscopically, to improve the surgical outcome of this clinical entity.