Role of LL-37 in Oral Bacterial DNA Accumulation in Dental Plaque.

Dental plaque, a highly structured polymicrobial biofilm, persistently forms in the oral cavity and is a common problem affecting oral health. The role of oral defense factors in either collaborating or disrupting host-microbiome interactions remains insufficiently elucidated. This study aims to explore the role of LL-37, a critical antimicrobial peptide in the oral cavity, in dental plaque formation. Through immunostaining dental plaque specimens, we observed that LL-37 and DNA colocalized in the samples, appearing as condensed clusters. In vitro experiments revealed that LL-37 binds rapidly to oral bacterial DNA, forming high molecular weight, DNase-resistant complexes. This interaction results in LL-37 losing its inherent antibacterial activity. Further, upon the addition of LL-37, we observed a visible increase in the precipitation of bacterial DNA. We also discovered a significant correlation between the levels of the DNA-LL-37 complex and LL-37 within dental plaque specimens, demonstrating the ubiquity of the complex within the biofilm. By using immunostaining on dental plaque specimens, we could determine that the DNA-LL-37 complex was present as condensed clusters and small bacterial cell-like structures. This suggests that LL-37 immediately associates with the released bacterial DNA to form complexes that subsequently diffuse. We also demonstrated that the complexes exhibited similar Toll-like receptor 9-stimulating activities across different bacterial species, including Porphyromonas gingivalis, Fusobacterium nucleatum, Prevotella intermedia, and Streptococcus salivarius. However, these complexes prompted dissimilar activities, such as the production of IL-1ß in monocytic cells via both NLRP3 pathway-dependent and pathway-independent mechanisms. This study, therefore, reveals the adverse role of LL-37 in dental plaque, where it binds bacterial DNA to form complexes that may precipitate to behave like an extracellular matrix. Furthermore, the unveiled stimulating properties and species-dependent activities of the oral bacterial DNA-LL-37 complexes enrich our understanding of dental plaque pathogenicity and periodontal innate immune responses.

Preserved Smad4 expression in the transforming growth factor beta signaling pathway is a favorable prognostic factor in patients with advanced gastric cancer.

The signals of the transforming growth factor beta (TGF-beta) superfamily are conveyed through cell surface serine/threonine kinase receptors to the intracellular mediators known as Smads. Activation of Smads causes their translocation from the cytoplasm to the nucleus, where they function to control gene expression. The present study analyzed the expression of Smad4 and TGF-beta1 to determine their prognostic significance in advanced gastric cancer. Of 249 cases of advanced gastric cancer, 41 had invaded the muscular layer, 114 had invaded the subserosal layer, and 94 had invaded the serosa. Anti-Smad4 and TGF-beta1 antibodies were used for immunohistochemical staining. Reduced expression of Smad4 was 75.1%, whereas positive expression of TGF-beta1 was 39.6% in gastric cancer. Smad4 expression was related to the depth of tumor invasion (P < 0.05), and TGF-beta1 expression correlated with tumor gross type (P < 0.05). Postoperative survival analysis indicated that patients who had a tumor with reduced Smad4 expression had a poorer clinical outcome than those with preserved expression (P < 0.05). Furthermore, in patients with TGF-beta1-positive tumors, survival rate was significantly better in patients with preserved Smad4 expression than in those with reduced Smad4 expression (P < 0.05). According to multivariate analysis, Smad4 expression acted as an independent prognostic factor. Smad4 expression, particularly in the TGF-beta pathway, is an effective predictor of outcome for patients with advanced gastric cancer.

p21 expression is a prognostic factor in patients with p53-negative gastric cancer.

The expression of p21 and p53 proteins was analyzed by immunohistochemistry in 256 patients with advanced gastric cancer. The results showed that strong, weak and negative expression of p21 were detected in 22.2 (57/256), 68.0 (174/256) and 9.8% (25/256) of the patients, respectively. p53 expression was found in 28.9% (74/256). The expression of p21 was not associated with clinicopathological features. In p53 negative tumors, p21 expression was associated with the survival of patients who underwent curative operations (P = 0.007). The 5-year survival rates were 20.1, 36.6 and 59.8% in patients with p21-negative, -weakly positive and -strongly positive tumors, respectively. In contrast, in p53-positive tumors, prognosis did not differ in spite of p21 expression. Multivariate analysis showed that p21 expression was an independent factor in patients with p53-negative tumors. These results indicate that examination of p21 expression in p53 negative tumors will be useful for estimating the prognosis of patients with advanced gastric cancer.

Selectins induced by interleukin-1beta on the human liver endothelial cells act as ligands for sialyl Lewis X-expressing human colon cancer cell metastasis.

We have previously reported that colon cancer cells metastasized to the liver expressed an increased amount of sialyl Lewis X (SLeX) antigen compared to their corresponding primary lesions. It is now well known that SLeX antigen and sialyl Lewis A (SLeA) antigen are ligands for the selectins expressed on the endothelial cells. Therefore, it is assumed that SLeX-rich colon cancer cells could be easily adhered to the endothelial cells that express selectins. In this report we have tried to induce selectin expression on the human liver sinusoidal endothelial cells and have examined the adhesion of SLeX-high or -low expressing colon cancer cells to the interleukin-1beta (IL-1beta)-treated liver specimens using Stamper-Woodruff assay. These human colon cancer cells are termed KM12HX or KM12LX cells, respectively. A significantly increased number of KM12HX cells adhered to the IL-1beta-treated liver specimens compared to KM12LX cells. The adhesion of KM12HX cells was inhibited by the pretreatment of tumor cells with anti-SLeX antibody or by the pretreatment of liver specimens with anti-selectin antibodies. Selectin expression on the liver sinusoidal endothelial cells and endothelial cells of blood vessels after IL-1beta treatment was confirmed by immunohistochemically using anti-selectin monoclonal antibodies (MAbs). These findings strongly suggest that SLeX-expressing cancer cells could adhere to the sinusoidal endothelial cells via an SLeX-selectin interaction system and this could be a first step for colon cancer cells that metastasize to the liver. The mechanism by which these selectins can be induced in vivo is the next problem to be considered.

Response of patients with cirrhosis who have undergone partial hepatectomy to treatment aimed at achieving supranormal oxygen delivery and consumption.

BACKGROUND: This study was undertaken to evaluate the response to therapy aimed at achieving supranormal cardiac and oxygen transport variables (cardiac index > than 4.5 L/min/m2, oxygen delivery > 600 ml/min/m2, and oxygen consumption > 170 ml/min/m2) in patients with cirrhosis who have undergone partial hepatectomy and to assess the relationship between those parameters and outcome. METHODS: Thirty-four consecutive patients underwent elective hepatectomy for hepatocellular carcinoma. The postoperative outcomes and hemodynamic and oxygen transport values in 16 patients (group S) who maintained supranormal values were compared with those in 18 patients (group N) treated to maintain normal hemodynamic values. Patients in group S received volume expansion and then, if necessary, dobutamine (3 to 15 micrograms/kg/min) to increase cardiac index, oxygen delivery, and oxygen comsumption simultaneously during the first 12 hours. RESULTS: The hemodynamic targets were reached by 56% of patients in group S during the first 12 hours and 31% during the next 12 hours. Postoperative blood lactate levels at 12 and 24 hours were lower in group S than in group N, and total bilirubin concentrations, hepatic venous oxygen saturation, and arterial ketone body ratio, useful markers of postoperative liver function, also showed more favorable changes in group S than in group N. Postoperative morbidity and mortality rates were not significantly different in the two groups, but the incidence of hyperbilirubinemia and liver failure was much lower in group S than in group N. CONCLUSIONS: These results suggest that fluid therapy aimed at achieving a supranormal pattern by 12 hours after hepatectomy improved the systemic oxygen demand-supply dynamics and hepatic hemodynamics, decreasing the incidence of postoperative hyperbilirubinemia and liver failure in patients with liver cirrhosis.

Exogenous hepatocyte growth factor markedly stimulates liver regeneration following portal branch ligation in dogs.

Portal branch ligation (PBL) or embolization prior to extensive hepatectomy has been employed to increase the functional reserve of the remaining liver. This study investigated the effects of human recombinant hepatocyte growth factor (rh-HGF) on liver regeneration following PBL in dogs. Beagle dogs were subjected to PBL and were divided into two groups, a control group (n = 11) without rh-HGF and a treated group (n = 12) receiving postoperative rh-HGF at 250 ng/kg via the portal vein. Dogs were killed 72 h or 14 days following PBL. We studied the changes in serum HGF level, DNA synthesis of the liver, hepatocyte size, liver weight, and liver function tests. In the HGF group, the ratio of whole liver weight to body weight increased significantly, and both ligated and nonligated lobes showed marked increases in weight. The nonligated lobes in the HGF group showed significant increases in both DNA synthesis and hepatocyte size. Moreover, ligated lobes in the HGF group showed an increase in DNA synthesis without hypertrophy compared with the control group. Administration of rh-HGF did not significantly affect liver function tests. Ligation of the portal branch supplying the portion of liver to be resected, coupled with the administration of rh-HGF, is a useful strategy to increase hepatic reserve in advance of major hepatectomy.

E-cadherin but not N-cadherin expression is correlated with the intracellular distribution of catenins in human hepatocellular carcinomas.

Cadherins are Ca2+-dependent cell-cell adhesion molecules, and are involved in the formation and maintenance of the histo-architecture. Using a combination of biochemical and immunohistochemical methods, we analyzed the expression of the cadherin-catenin complex in 34 human hepatocellular carcinomas. Unexpectedly, we found the expression of N (neural)-cadherin in normal hepatocytes and all hepatocellular carcinomas examined. In 18 cases, the decreased expression of E (epithelial)-cadherin was observed. Among them, the decreased expression of alpha-catenin and gamma-catenin (plakoglobin) was also observed in 9 and 6 cases, respectively. Thus the decreased expression of alpha-catenin and gamma-catenin was apparently preceded by the decreased expression of E-cadherin. The decreased expression of beta-catenin was not observed in any of the cases analyzed. beta-Catenin was found to accumulate in the cytoplasm of hepatocellular carcinomas with the decreased expression of E-cadherin, despite the presence of N-cadherin at the cell-cell contacts. These results suggest a pivotal role of E-cadherin in the intracellular distribution of catenins in hepatocellular carcinomas.

[A study of MTT-hybrid assay using human bone and soft tissue tumor cells].

We investigated a new chemosensitivity test, MTT-hybrid assay, which was a hybrid of MTT colorimetric assay and double-layered soft agar colony assay, using human bone and soft tissue tumor cells. MTT formazan crystals produced by viable cells in the soft agar medium were solubilized by SDS at 60 degrees C. The absorbance (560 nm) is directly proportional to the cell number over a wide range. The absorbance increased in proportion to colonial growth of osteosarcoma cells, while it decreased in a human diploid cell strain in a few days. Drug sensitivity of tumor cells is supposed to be assessed without contaminating normal cells by MTT-hybrid assay in primary tumor samples. Good correlation of IC50 was observed between MTT-hybrid assay and colony assay. The MTT-hybrid assay shows potential value as a rapid predictive test for chemotherapeutic agents in an individual patient.

Surgical treatment of cervical spondylotic myelopathy complicating athetoid cerebral palsy.

Operative treatment was performed in nine patients with cervical spondylotic myelopathy complicating athetoid cerebral palsy. The first two patients were treated by laminectomy, and the other seven by anterior interbody fusion. The symptoms in both the laminectomy patients improved after operation, but became worse again when cervical instability developed; they then had to have an anterior fusion in addition. In six of the seven patients who had primary anterior fusion a halo-cast (or a halo-vest) was used to keep the cervical spine immobile, and good bony fusion was obtained with satisfactory results. However, in one patient no halo apparatus was used, bony union did not occur and the radiculopathy reappeared. In cervical myelopathy complicating athetoid cerebral palsy laminectomy is contra-indicated; anterior fusion combined with a halo apparatus is, however, satisfactory.

Early post-operative hyperbaric oxygen therapy modifies neutrophile activation.

BACKGROUND/AIMS: To investigate the effect of acute hyperbaric oxygen therapy (HBOT) on post-operative sinusoidal endothelial cell (SEC) damage caused by activated neutrophils. METHODOLOGY: 12 non-cirrhotic patients (Group H), who underwent elective hepatectomy for liver cancer, were given 2 courses of HBOT: 2.0 atm with inhalation of 100% oxygen, for 60 min, at 3 hours and 24 hours after hepatectomy; they were then compared with the 12 patients (Group C) who had been treated to maintain normal hemodynamic values. RESULTS: In group H, peak levels of polymorphonuclear leukocyte elastase (PMNE) and thrombomodulin (TM) were clearly diminished and delayed compared to Group C. All subjects in Group C showed more than a 10% increase in CD18 12 hours after surgery; however, in Group H, the elevation of CD18 expression was clearly suppressed compared to Group C. No patient in Group H had post-operative hyperbilirubinemia or hepatic failure; however, 3 had post-operative hyperbilirubinemia and 1 had intraperitoneal infection in Group C. CONCLUSIONS: Our results provide direct evidence that HBOT, especially at 3 hours after hepatectomy, has favorable effects on the activation of neutrophiles decreasing SEC injury.

Risk prediction using histology of noncancerous liver before hepatic resection for hepatocellular carcinoma.

BACKGROUND/AIMS: The aim of this study is to elucidate the feasibility of the risk assessment of hepatic resection by histological evaluation of noncancerous liver in patients with hepatocellular carcinoma. METHODOLOGY: The study involved 78 patients with hepatocellular carcinoma who had undergone a needle biopsy of noncancerous liver before hepatic resection. The histological activity index score which consists of four categories indicating the inflammatory activity and the degree of fibrosis was determined, and its association with complications after hepatic resection was examined. RESULTS: Postoperative complications occurred in 26 of the first 52 patients that underwent hepatic resection. A logistic analysis selected histological activity index score as an independent factor related to postoperative complications (Odds ratio 1.31, P < 0.02). Postoperative complications occurred more frequently in patients with a histological activity index score > or = 6 that had undergone resection of two or more segments (P < 0.05), and also in those with histological activity index score > or = 10 that had undergone segmentectomy or subsegmentectomy (P < 0.05). When the histological activity index score was taken into consideration in deciding operative procedures for a further 20 patients, the incidence of postoperative complications reduced considerably to 10%. CONCLUSIONS: Preoperative histological evaluation of noncancerous liver by a needle biopsy may be helpful in deciding the operative procedure to avoid complications after hepatic resection for hepatocellular carcinoma.

Hepatocyte growth factor promotes liver regeneration and protein synthesis after hepatectomy in cirrhotic rats.

BACKGROUND/AIMS: Hepatocyte growth factor, a potent mitogen for hepatocytes has been reported to be a hepatrophic factor in normal livers. In this study, the effect of exogenous hepatocyte growth factor on liver regeneration in cirrhotic rats was investigated, in vitro and in vivo. METHODOLOGY: Liver cirrhosis was induced by intraperitoneal injections of an emulsion, carbon tetrachloride and olive oil, twice weekly for 10 weeks. In vitro, various amounts of exogenous hepatocyte growth factor; 0, 0.5, 1, 2.5, 5, and 10 ng/mL; were added to the hepatocytes isolated using in situ perfusion method. In vivo, partial hepatectomy (Hx), according to the procedure described by Higgins and Anderson, was performed on cirrhotic rats. Saline solution (control group) or 3 micrograms/kg of exogenous hepatocyte growth factor (HGF group) was then injected through the tail vein at intervals 12 hours after Hx. RESULTS: In vitro, DNA synthesis in hepatocytes obtained from cirrhotic livers increased following exogenous hepatocyte growth factor in dose-dependent fashion. In vivo, the labeling index of 5-bromo-2'-deoxyuridine at 24 hours after Hx was markedly increased by exogenous hepatocyte growth factor (control, 10.0 +/- 3.1%; hepatocyte growth factor, 25.8 +/- 9.8%; P < 0.01). Furthermore, serum albumin at 24 and 72 hours and a normotest at 24 hours after Hx, were significantly higher in the HGF group than in the control group. CONCLUSIONS: These results indicate that exogenous hepatocyte growth factor may promote DNA synthesis and protein synthesis during liver regeneration after Hx with cirrhosis.

Favorable quality of life after repeat hepatic resection for recurrent hepatocellular carcinoma.

BACKGROUND/AIMS: The appropriate choice of treatment for recurrent hepatocellular carcinoma after hepatic resection remains controversial. The aim of this study is to clarify prognostic factors and quality of life in patients with tumor recurrence after hepatic resection for hepatocellular carcinoma. METHODOLOGY: We retrospectively analyzed 188 patients with hepatocellular carcinoma who underwent curative hepatic resection between 1988 and 1997. Statistical analysis was performed to identify prognostic factors involved after recurrence. Furthermore, quality of life after treatment for recurrence was compared between patients with repeat hepatic resection or hepatic arterial infusion chemotherapy. RESULTS: In 123 patients with recurrence, unfavorable predictors after recurrence are pTNM Stage III/IV at initial surgery, receiving chemotherapy before initial surgery and presence of extrahepatic recurrence. In contrast, favorable predictors are 3 years or more of disease-free interval and repeat hepatic resection. The incidence of deteriorated performance status in the repeat hepatic resection group was lower than in the hepatic arterial infusion chemotherapy group because of better psychological function in patients undergoing repeat hepatic resection. CONCLUSIONS: Repeat hepatic resection provides a good prognosis and a favorable quality of life in patients with recurrence after hepatic resection for hepatocellular carcinoma.

A comparison of hepatocellular carcinoma associated with HBV or HCV infection.

BACKGROUND/AIMS: Close relationships between hepatocellular carcinoma (HCC) and hepatitis virus infection have been elucidated. However, clinical differences between HBV- and HCV-associated HCC remain unclear. METHODOLOGY: Four hundred and sixteen patients with HCC were examined for both HBsAg and HCV-Ab. Sixty-nine of the 416 patients (16.6%) were positive for HBsAg and negative for HCV-Ab (B-HCC), while 290 patients (69.7%) were negative for HBsAg and positive for HCV-Ab (C-HCC). Various comparisons were made between the 2 groups. RESULTS: B-HCC patients were younger in age (p < 0.0001), and had significantly better liver function than C-HCC patients. The motivation of HCC detection was different between the 2 groups (p < 0.0001), and the tumor size of B-HCC was significantly larger when comparing groups with regard to symptoms (p < 0.05). Although B-HCC demonstrated better survival in Stage I/II treated by surgery (p < 0.05), it was associated with poorer survival in Stage III/IV receiving hepatic arterial infusion chemotherapy when compared to C-HCC (p < 0.01). CONCLUSIONS: These results suggest that clinical differences between B-HCC and C-HCC may depend upon the difference of the natural course between HBV and HCV infection, and B-HCC may be more resistant to treatment than C-HCC in the advanced stage. This also illustrates the need for early tumor detection in B-HCC.

Operative treatment of congenital musclar torticollis: long term results of myotomy.

Myotomy was performed in 40 patients under 10 years of age with typical congenital muscular torticollis. Follow-up studies were carried out on 29 out of 40 cases. The patients without muscle relief of the affected sternomastoid muscle after myotomy had good range of motion of the neck. On the other hand, the patients who had undergone muscle release had, to grater or lesser extent, disturbances of the range of movement. For complete release of contracture, total excision may be the only operation in younger cases.

A new open reduction treatment for congenital hip dislocation: long-term follow-up of the extensive anterolateral approach.

Congenital hip dislocation, which is conservatively unmanageable, has usually been treated using open reduction. However, a long-term follow-up study of the results suggests that this procedure is unsatisfactory. Since 1973, Tanabe has used a new open reduction procedure that circumferentially dissects the joint capsule and produces sufficient concentric reduction of the femoral head in the acetabulum immediately after the surgery. Fifty-six children (65 hips) from the age of 1 to 3 years were treated by this procedure, and fifty-one of them were clinically and roentgenographically followed up from 6.3 to 12.4 years after the surgery. At the final follow-up session, all children had grown to be over 9 years of age, and no patient had clinically significant symptoms. According to Severin's classification, 33 hips were rated in Group I, and 14 hips in Group II. Another 10 hips were in Group III, and one hip was in Group IV. The incidence of avascular necrosis was 5.2 per cent. These data suggest that our procedure is more useful than the previous ones.

Unique features of monoclonal IgG2b in the cleavage reaction with pepsin.

Preparations of IgG2b purified from several mouse hybridoma clones were highly susceptible, compared to other subclasses, to peptic digestion under conditions usually used to prepare F (ab')2 fragments. Analyses of the digestion products revealed that no F (ab')2 was produced and that the main product was a Fab-like fragment. Demonstration of the hinge disulfides in the Fc portion clearly indicated that in IgG2b the primary peptic cleavage occurs on the NH2-terminal side of the inter-heavy chain disulfide bridge. The resulting Fab failed to bind with antigen, suggesting the importance of the CH1-hinge region in maintaining the native conformation of the antigen-binding site.

Tumor angiogenesis related to growth pattern and lymph node metastasis in early gastric cancer.

OBJECTIVE: To investigate the correlation between angiogenesis and tumor growth pattern as well as the lymph node metastasis to reveal the significance of vascularity in the early stage of gastric cancer. METHODS: 97 specimens from patients with early gastric cancer were studied by immunohistochemical method using anti-factor VIII related antigen antibody. RESULTS: Microvessel count was related to tumor growth pattern. The mean vessel count was higher in superficially spreading and penetrating types of tumors. Lymph node metastasis was correlated to microvessel count. Tumors with lymph node metastasis had higher microvessel counts than those without lymph node metastasis. CONCLUSIONS: In the early stage of gastric carcinoma, angiogenesis is correlated with tumor growth pattern and lymph node metastasis. Identification of tumors with high density of vascularization is beneficial for closer follow-up and adjuvant therapy.

[Clinicopathological studies of the recurrence of esophageal squamous cell cancer--with special reference to the mode of recurrence and operative procedure].

In 93 out of 201 patients (46%) with squamous cell carcinoma of the esophagus who underwent radical resection (excluding death within 30 days after operation), the site of recurrence could be identified by means of X-ray, CT, ultrasonography, and biopsy. Recurrence was found in 55% of 93 cases within 12 months after surgery and in 86% of 93 cases within 24 months. Of 93 patients with recurrences, lymph node recurrences were present in 44 cases, visceral recurrences in 32 cases, both lymph node and visceral recurrences in 11 cases and others in 6 cases. Neck and/or upper mediastinal lymph node recurrences were found in 10 out of 15 patients who had recurrences within 3 months after surgery. Careful examination should be made in the left recurrent nerve chain and extended lymph node resection of upper mediastinal region should be performed under the adequate indication. Esophageal squamous cell cancer has a tendency to recur in the lymph nodes initially, and visceral metastases may occur thereafter. The incidence of visceral recurrence increased remarkably, when neck and/or upper mediastinal lymph nodes were involved at the time of operation. Accordingly, both irradiation and chemotherapy should also be applied for improving the prognosis of esophageal carcinoma.

[Pre-operative staging in advanced gastric cancer by ultrasound; with special reference to para-aortic lymph node metastases].

Extraluminal cancer spreadings such as peritoneal dissemination, liver metastases, lymph node metastases and infiltration to surrounding organs were examined in 236 patients with gastric cancer by ultrasound (US). Peritoneal dissemination was found in 36 of 236 cases. Ascites, nodules in the cul-de-sac and thickened wall of the transverse colon could be detected by US in 24 of 36 cases (sensitivity of 67%). Liver metastases were detected in 18 out of 23 cases (78%). Infiltration to the surrounding organs, mainly to the pancreas, was impossible to demonstrate by US. Sensitivity was 44% (17/39). Lymph node metastases in the region around the celiac axis were indicated in 30 of 40 cases (75%) by US. Para-aortic nodal metastases were also detected in 27 of 34 cases (79%). Ultrasonic images of para-aortic lymph node metastases were classified into four figures: plate type, sandwich type, unilateral multiple type, unilateral solitary type. These types were not necessarily the same in indication of laparotomy, selection of combined or adjuvant therapy, significance of nodal dissection and also prognosis. Solitary involved para-aortic node near the left renal vein should be removed in a case free from peritoneal dissemination or liver metastases. Ultrasound could be very useful in screening patients with gastric cancer for peritoneal dissemination, liver metastases and lymph node metastases.