Investigation of the regeneration potential of the recurrent laryngeal nerve (RLN) after compression injury, using neuromonitoring.

INTRODUCTION: The aim of this study was to investigate the regeneration potential of RLN after the compression of the nerve, without disrupting its continuity, using neuromonitoring. METHODS: In the first operation, the RLN and nervus vagus of adult Goettingen minipigs were dissected free, and the neuromonitoring parameters (amplitude, threshold and lag time of signal) were measured. Injury of the RLN was induced using a "bulldog" clamp. When the signal was no longer detectable, after the 15 min regeneration phase, the operation was finished. The neuromonitoring studies (see above) were repeated in a second operation 6 months later. RESULTS: (1) After the first operation, acute clamping of the RLN led to a reduction in the amplitude of the neuromonitoring signal; the lag time and the threshold of signal remained. Complete restitution of the signal was observed during the first regeneration phase. Repeated clamping led to complete disappearance of the signal. (2) During the second operation, i.e., after 6 months of regeneration, the neuromonitoring signals of both RLN and nervus vagus were detected in 93% of the GMP. No statistical differences (p = 0.17) were noticed between the amplitude of the RLN before the nerve injury (first operation) and after nerve regeneration (second operation). A significant increase in the lag time (p < 0.0005) was shown for both RLN and nervus vagus. CONCLUSIONS: The acute compression of RLN can only be detected by observing the amplitude of the neuromonitoring signal. Restitutio ad integrum is possible after a short clamping period but it is important to preserve the RLN continuity.

The alternative sigma factor sigma B of Staphylococcus aureus modulates virulence in experimental central venous catheter-related infections.

The impact of the alternative sigma factor sigma B (SigB) on pathogenesis of Staphylococcus aureus is not conclusively clarified. In this study, a central venous catheter (CVC) related model of multiorgan infection was used to investigate the role of SigB for the pathogenesis of S. aureus infections and biofilm formation in vivo. Analysis of two SigB-positive wild-type strains and their isogenic mutants revealed uniformly that the wild-type was significantly more virulent than the SigB-deficient mutant. The observed difference in virulence was apparently not linked to the capability of the strains to form biofilms in vivo since wild-type and mutant strains were able to produce biofilm layers inside of the catheter. The data strongly indicate that the alternative sigma factor SigB plays a role in CVC-associated infections caused by S. aureus.

Long-term benefits of Roux-en-Y pouch reconstruction after total gastrectomy: a randomized trial.

OBJECTIVE: Roux-en-Y reconstruction with and without jejunal pouch was compared in a randomized controlled trial to identify the optimal reconstruction procedure in terms of quality of life. BACKGROUND DATA: Randomized trials comparing techniques of reconstruction after total gastrectomy have shown controversial results. METHODS: One hundred and thirty-eight patients with gastric cancer were intraoperatively randomized for Roux-en-Y reconstruction with pouch (n = 71) or without pouch (n = 67) after gastrectomy and stratified into curative or palliative resection. Intra- and postoperative complications were recorded. Body weight and quality of life were determined every 6 months with a follow-up of up to 12 years. RESULTS: Both groups were comparable for age, sex, incidence of concomitant disease, and staging. There were no differences in operative time, postoperative complications, and mortality. Short- and long-term weight loss was similar in both groups. In the first postoperative year, there were no benefits of pouch reconstruction in terms of quality of life, independent of the resection status. In the third, fourth, and fifth year after surgery quality of life was significantly improved for patients with a pouch. CONCLUSIONS: Roux-en-Y pouch reconstruction after gastrectomy is simple to perform and safe. Long-term survivors benefit from pouch reconstruction. Therefore, a pouch is recommended for patients with a good prognosis.

Growth of human gastric cancer cells in nude mice is delayed by a ketogenic diet supplemented with omega-3 fatty acids and medium-chain triglycerides.

BACKGROUND: Among the most prominent metabolic alterations in cancer cells are the increase in glucose consumption and the conversion of glucose to lactic acid via the reduction of pyruvate even in the presence of oxygen. This phenomenon, known as aerobic glycolysis or the Warburg effect, may provide a rationale for therapeutic strategies that inhibit tumour growth by administration of a ketogenic diet with average protein but low in carbohydrates and high in fat enriched with omega-3 fatty acids and medium-chain triglycerides (MCT). METHODS: Twenty-four female NMRI nude mice were injected subcutaneously with tumour cells of the gastric adenocarcinoma cell line 23132/87. The animals were then randomly split into two feeding groups and fed either a ketogenic diet (KD group; n = 12) or a standard diet (SD group; n = 12) ad libitum. Experiments were ended upon attainment of the target tumor volume of 600 mm3 to 700 mm3. The two diets were compared based on tumour growth and survival time (interval between tumour cell injection and attainment of target tumour volume). RESULTS: The ketogenic diet was well accepted by the KD mice. The tumour growth in the KD group was significantly delayed compared to that in the SD group. Tumours in the KD group reached the target tumour volume at 34.2 +/- 8.5 days versus only 23.3 +/- 3.9 days in the SD group. After day 20, tumours in the KD group grew faster although the differences in mean tumour growth continued significantly. Importantly, they revealed significantly larger necrotic areas than tumours of the SD group and the areas with vital tumour cells appear to have had fewer vessels than tumours of the SD group. Viable tumour cells in the border zone surrounding the necrotic areas of tumours of both groups exhibited a glycolytic phenotype with expression of glucose transporter-1 and transketolase-like 1 enzyme. CONCLUSION: Application of an unrestricted ketogenic diet enriched with omega-3 fatty acids and MCT delayed tumour growth in a mouse xenograft model. Further studies are needed to address the impact of this diet on other tumour-relevant functions such as invasive growth and metastasis.

Neoadjuvant therapy of gastric cancer with the human monoclonal IgM antibody SC-1: impact on the immune system.

Adjuvant therapies for minimal residual disease are a promising approach to improve the poor survival rates after surgery of gastric tumors. A pilot study of a neoadjuvant therapy was performed using a human monoclonal IgM antibody (SC-1) specifically inducing apoptosis in signet ring cell stomach carcinomas. However, scarce information exists on how such a treatment affects the immune system, in particular what are the effects of apoptosis induction and infusion of large amounts of IgM. Thus, the leukocyte composition (CD3, CD4, CD8, CD19, CD16+56, CD14) and several cytokines (TNF-alpha, IL6, IL12, IFN-gamma, GM-CSF, Neopterin) before and after SC-1 application were measured and compared to results of patients that underwent surgical removal of gastric carcinoma without antibody treatment. After SC-1 application, an increase in TNF-alpha and a decrease of lymphocytes and CD3+ T-cells but in the range obtained in healthy individuals was observed before surgery. After surgery, the IL6 levels increased and the TNF-alpha levels remained at the elevated level. Furthermore, there was a significant drop in lymphocytes and CD3+ T-cells. These effects were due to the surgical treatment. Other parameters did not show significant changes. It seems that the application of an apoptosis-inducing antibody prior to surgery of gastric tumors has mild if any effect on the immune system. Therefore, from an immunological point of view, the treatment with this monoclonal antibody is extremely safe.

Severe pyomyositis caused by Panton-Valentine leucocidin-positive methicillin-sensitive Staphylococcus aureus complicating a pilonidal cyst.

BACKGROUND: To our best knowledge, Panton-Valentine leucocidin (PVL)-positive methicillin-sensitive Staphylococcus aureus (MSSA) has not been described yet as cause for severe pyomyositis. CASE REPORT: We present a 23-year-old apparently healthy male patient without any typical predisposing findings who developed severe pyomyositis secondary to an operated pilonidal cyst. In the follow-up, the patient showed signs of immunocompromisation. The causative agent for purulent infection of multiple muscles was a MSSA strain harbouring PVL toxin. RESULTS: In the reported case, aggressive antibiotic and surgical treatment with additional application of immunoglobulins has lead to recovery from the disease without relapse. CONCLUSIONS: PVL-positive S. aureus are associated with skin diseases, multiple abscesses and often complicated by severe sepsis and necrotising pneumonia. Under such circumstances, the mortality rate can reach up to 75%. In addition, the PVL toxin can cause immunocompromisation and might be therefore involved in the aetiology of pyomyositis. Aggressive antibiotic and surgical treatment with additional application of immunoglobulins is recommended for treatment.

T-cell response to p53 tumor-associated antigen in patients with colorectal carcinoma.

Despite the radical surgical resection performed in patients with colorectal carcinoma, there is a high rate of tumor recurrence. Over an observation period of 3 years, 18% of the patients in our collective suffered a tumor relapse with local or distinct metastases after initial R0-resection. Some evidence suggests that this may be due to suppression of anti-tumor responses, a phenomenon that might be attributed to regulatory T cells. The aim of our study was to investigate the tumor-specific immune response depending on the UICC stage of patients with colorectal cancer. The cellular immune responses against defined antigens that are overexpressed in most of the patients with colorectal cancer were characterized. For this purpose, the tumor suppressor gene, p53, was chosen as the tumor-associated antigen that exhibits mutations and overexpression in up to 60% of colorectal carcinoma. We observed that p53 induced both IFN-gamma and IL-10 secretion. The predominance of IL-10 production indicated that regulatory T cells directly participate in modulating the anti-tumor immune response. IL-10 levels in the blood as well as the expression of regulatory T-cell specific genes at the tumor site correlate with the UICC stage of the disease. These results may provide an explanation for the poor prognosis and increased recurrence rate in patients with advanced carcinoma.

Feasibility and limits of an orthotopic human colon cancer model in nude mice.

We sought to develop an accurate colorectal cancer model in nude mice with stable local growth, tumor cell dissemination, and reproducible metastatic capacity. To this end, we orthotopically transplanted histologically intact human colorectal cancer tissue from 10 human patients into nude mice. After successful local tumor growth, tumor tissues were retransplanted as many as 9 times in serial passage. All specimens were transplanted using microsurgical techniques. Histologic, immunohistochemical, and polymerase chain reaction techniques were used to determine tumor growth rates and kinetics, development of regional lymph node and distant hepatic metastases, and the induction of minimal residual disease (MRD). Stable local tumor growth rates with variable growth kinetics were detected in 73.4% of all mice. The lymph node and hepatic metastasis rates were low, at 18.4% and 4.9%, respectively. MRD, as reflected by CK20 positivity of the bone marrow in animals with lymph node and hepatic metastases, was present in 22.2%. The orthotopic colorectal cancer model described here is feasible for the induction of reproducible local tumor growth but is limited by variable growth kinetics and the low rate of lymph node and hepatic metastases. Cytokeratin-positive cells indicative of MRD could be detected in the bone marrow of approximately 25% of the nude mice with metastases. The observed induction of MRD after orthotopic implantation of intact human colon cancer in animals with lymph node and hepatic metastases might be improved if established colon cancer cell lines were used.

[Extract of Passiflora edulis in the healing process of gastric sutures in rats: a morphological and tensiometric study]. / Efeito do extrato da Passiflora edulis na cicatrização de gastrorrafias em ratos: estudo morfológico e tensiométrico.

INTRODUCTION: Many substances of vegetable origin have been used since the beginning of civilization with the purpose of improving the healing process. Among them, dry leaves extract from Passiflora edulis have been shown to have an anti-inflammatory effect in rats. PURPOSE: To analyze the effect of dry leaves extract from Passiflora edulis in the healing of gastric sutures in rats. METHODS: Forty male adult Wistar rats were divided into two groups of 20 rats, called Passiflora group (GP) and Control group (GC) which were divided into two groups of 10 according to moment of death, on day 3 or day 7 after the operation. All animals were submitted to a midline incision and a gastrotomy was performed on the anterior wall of the stomach followed by gastric suture with polypropylene 6.0 using four stitches on a single layer. Rats from GP were given, before closure of the abdominal wall, a solution of Passiflora edulis extract, 250 mg/kg/weight, while rats from the GC were given an isovolumetric isotonic saline solution. Macroscopic evaluation included the adhesion index proposed by Knightly. Bursting pressure was measure by an electronic device. Microscopic analysis was performed including inflammation parameters. RESULTS: All animals presented adequate healing of the abdominal wall with no clinical signs of infections or dehiscence. The adherence index was similar in both groups both on day 3 (p=0.734) and on day 7 (p=1.000). The gastric sutures presented leak with smaller insufflation pressure on the 3rd P.O. day in both groups as compared to the 3rd P.O. day. There was no significant difference of bursting pressure among the subgroups on the 3rd P.O. day (GC3=41.1 +/- 22.1 mmHg versus GP3=59.2 +/- 20.4 mmHg; p=0.074). On the 7th P.O. day, there was an increased mean bursting pressure in both groups, but there was no statistically significant difference between the two groups (p=0.850). Histologic parameters were similar in both groups, on P.O. days 3 and day 7, except for the fibroblastic proliferation, which was greater on the 7th day in GP (p=0.002). CONCLUSION: The intraperitoneal use of Passiflora edulis extract influences favorably the healing of gastric sutures in rats because of the increase in the fibroblastic proliferation on the 7th P.O. day.

[Effect of Passiflora edulis (passion fruit) extract on rats' bladder wound healing: morphological study]. / Efeito do extrato de Passiflora edulis (maracujá) na cicatrização de bexiga em ratos: estudo morfológico.

PURPOSE: To evaluate the effects of hydroalcoholic extract of Passiflora edulis leaves in the healing of urinary bladder in rats from histological aspects. METHODS: Forty Wistar male rats were submitted to a longitudinal incision of the bladder followed by a stetching in only one level. After this common procedure, animals were divided at random two groups: Passiflora and Control. In the Passiflora group the only dosage used was administered by intraperitoneal injection of hydroalcoholic extract of Passiflora edulis leaves while in the Control group distilled water was injected. Each subgroup was then divided in two subgroups according to the death of these animals: Control, three and seven days, Passiflora, three and seven days. After the death of these animals, an inventory of the abdominal cavity was performed and the bladder was removed. A comparative analysis was done between the two groups with microscopic evaluation of the healing. There was less acute inflammation (p=0.008), greater collagenous formation (p=0.001) and greater capillary neo-formation (p=0.000) in the third day Passiflora subgroup when compared to the Control subgroup of the third day. RESULTS: There was less acute inflammation (p=0.001), greater fibroblastic proliferation (p=0.011) and greater collagenous formation (p=0.001) in the Passiflora subgroup of seventh day when compared with the Control seventh day subgroup. CONCLUSION: The use of Passiflora edulis leaves extract resulted in less acute inflammation, greater fibroblastic proliferation, collagenous formation and capillary neo-formation on rats' bladder wound healing.

Human antibody SC-1 reduces disseminated tumor cells in nude mice with human gastric cancer.

Advanced gastric cancer is a systemic disease that requires adjuvant therapy targeted at eliminating disseminated tumor cells (DTCs). We investigated whether the apoptosis-inducing human monoclonal IgM antibody SC-1 was able to reduce the number of disseminated gastric cancer cells in blood and bone marrow. Human gastric tumor specimens with positive expression of the SC-1 receptor were transplanted in nude mice with metastasizing gastric cancer. After tumor growth (4-6 weeks) animals were randomly allocated to intraperitoneal 100 microg SC-1 (n=23) or 100 microg human IgM (n=23). One week later, animals were sacrificed and blood and bone marrow specimens were obtained. A nested RT-PCR for cytokeratin 20 (CK-20) from blood and bone marrow of mice was performed for detection of disseminated tumor cells. Animals receiving SC-1 had significantly fewer DTCs than did control animals (p=0.0011). None of the SC-1 mice had DTCs simultaneously in both blood and bone marrow versus four of the control animals (p=0.0363). The reduction of DTCs in SC-1 animals was due to reduction in bone marrow (p=0.032 compared to controls), but not in blood (p=0.1158). Treatment with SC-1 significantly reduced the number of DTCs in bone marrow in this animal model.

Capillary activity of surgical sutures and suture-dependent bacterial transport: a qualitative study.

OBJECTIVE: To evaluate the multitude of new synthetic absorbable sutures (both monofilament and multifilament) in comparison with older materials with regard to capillarity and bacterial transport. METHODS: Sutures of United States Pharmacopoeia (USP) 4-0 thickness were arranged in a three-chamber system under sterile conditions. Either a colorant (liquid transport evaluation) or bacteria (bacterial transport evaluation) were added to the contamination chamber, and movement of colorant or bacteria was evaluated for as long as 30 days. RESULTS: None of the monofilament sutures transported colorant or bacteria. Colorant transport was found on the pseudomonofilament and multifilament sutures between the first and the fifth day. Escherichia coli were transported on the majority of the multifilament sutures, although no transport was found on silk or polyester sutures. Bacterial transport was most often evident in tests using the motile Proteus mirabilis. CONCLUSIONS: All multifilament and pseudomonofilament suture designs allowed transport of colorants and bacteria to some degree. The movement of fluids and bacteria did not depend on the absorptive capacity of the sutures, coating, or the presence of an open suture end.

Detection of disseminated tumor cells in nude mice with human gastric cancer.

Disseminated tumor cells (DTC) are a potential contributor to relapse of cancer. In the present study we developed a model for induction of disseminated tumor cells in nude mice, which can aid in the search for therapeutic approaches as well as improve our understanding of metastasizing gastric cancer. To detect DTC in blood and bone marrow we established a modified animal model of orthotopic transplantation. Two groups of nude mice were used for xenotransplantation of gastric cancer specimens. In group I tumor specimens originating from a gastric adenocarcinoma cell line were transplanted onto the stomach; in group II they were transplanted subcutaneously into both axillaries. Tumor growth, metastases and presence of DTC were compared in both groups. For detection of DTC a nested reverse transcriptase polymerase chain reaction (RT-PCR) for human cytokeratin (CK)-20 was performed on blood and bone marrow of all mice. Tumor growth occurred in both groups (9/10 animals in group I, 10/10 in group II) within 14 weeks. Only animals in group I developed local invasive tumor growth, stenosis of the stomach and distant metastases. Tumors in animals of group II grew with local displacement only and developed no metastases. There were no signals of CK-20 detected in the blood in both groups. In group I, 5 of 9 animals had positive signals of human CK-20 in their bone marrow as a sign of DTC. In group II no DTC were detected in bone marrow. We conclude that orthotopic transplantation is a prerequisite for the development of DTC and metastasizing tumor growth in this modified gastric cancer model.

Monoclonal antibody against beta7 integrins, but not beta7 deficiency, attenuates intestinal allograft rejection in mice.

BACKGROUND: beta7 integrins mediate homing and retention of lymphocytes to the normal and inflamed small bowel in a tissue-selective fashion. In the present study, we investigated the expression of beta7 integrins after small bowel transplantation (SBT) and tested the effects of blocking beta7-mediated pathways by using monoclonal antibody (mAb) or knockout mice. METHODS: Heterotopic SBT from BALB/c to C57BL/6 (B6) was used as a surgical model. Expression of beta7 integrins was measured on recipient lymphocytes (CD4 and CD8 ) in spleen, blood, and mesenteric lymph nodes (MLN) using flow cytometry. To analyze the effects of blocking beta7 integrins on graft survival, either beta7-deficient B6 or wild-type B6 mice that were treated with mAb against beta7 were studied. RESULTS: After allogeneic SBT, there were markedly increased levels of alpha4beta7 recipient CD8 lymphocytes in MLN, blood, and spleen as early as 3 days postoperatively. In contrast, alpha4beta7 integrin levels in isograft recipients were similar to those of normal mice. Mean survival time of intestinal allografts was not affected by beta7 deficiency (7.3+/-1 days) compared with wild-type mice (7.5+/-0.8 days). However, mAb against beta7 integrins significantly prolonged graft survival (12.8+/-1 days) compared with treatment with control mAb (7.3+/-1 days, <0.001). Histologic changes of SBT rejection were significantly attenuated when mice were given mAb against beta7. CONCLUSION: As indicated by the increased levels of alpha4beta7 CD8 lymphocytes, activation of this integrin contributes to the immune response in SBT rejection. Furthermore, blocking beta7 integrins with mAb provides a suitable target for immunosuppressive therapy. The discrepancy in survival data obtained by mAb and beta7 deficiency may be a result of the more rapid activation of compensatory mechanisms in the knockout mice.