Prediction of pouchitis after ileal pouch-anal anastomosis in patients with ulcerative colitis using artificial intelligence and deep learning.

BACKGROUND: Pouchitis is one of the major postoperative complications of ulcerative colitis (UC), and it is still difficult to predict the development of pouchitis after ileal pouch-anal anastomosis (IPAA) in UC patients. In this study, we examined whether a deep learning (DL) model could predict the development of pouchitis. METHODS: UC patients who underwent two-stage restorative proctocolectomy with IPAA at Keio University Hospital were included in this retrospective analysis. The modified pouchitis disease activity index (mPDAI) was evaluated by the clinical and endoscopic findings. Pouchitis was defined as an mPDAI ≥ 5.860; endoscopic pouch images before ileostomy closure were collected. A convolutional neural network was used as the DL model, and the prediction rates of pouchitis after ileostomy closure were evaluated by fivefold cross-validation. RESULTS: A total of 43 patients were included (24 males and 19 females, mean age 39.2 ± 13.2 years). Pouchitis occurred in 14 (33%) patients after ileostomy closure. In less than half of the patients, mPDAI scores matched before and after ileostomy closure. Most of patients whose mPDAI scores did not match before and after ileostomy closure had worse mPDAI scores after than before. The prediction rate of pouchitis calculated by the area under the curve using the DL model was 84%. Conversely, the prediction rate of pouchitis using mPDAI before ileostomy closure was 62%. CONCLUSION: The prediction rate of pouchitis using the DL model was more than 20% higher than that using mPDAI, suggesting the utility of the DL model as a prediction model for the development of pouchitis. It could also be used to determine early interventions for pouchitis.

Smoking is a risk factor for pulmonary metastasis in colorectal cancer.

AIM: The hepatic microenvironment, which may include chronic inflammation and fibrosis, is considered to contribute to the pathogenesis of liver metastases of colorectal cancer. A similar mechanism is anticipated for pulmonary metastases, although no reports are available. Smoking causes pulmonary inflammation and fibrosis. Thus, we hypothesized that smokers would be especially affected by pulmonary metastases of colorectal cancer. In this study, we attempted to clarify the impact of smoking on pulmonary metastasis of colorectal cancer. METHOD: Between September 2005 and December 2010 we reviewed 567 patients with pathological Stage I, II or III colorectal cancer, whose clinicopathological background included a preoperative smoking history, pack-year history from medical records. Univariate and multivariate analyses using the Cox proportional hazard model were performed to determine the independent prognostic factors for pulmonary metastasis-free survival. RESULTS: Pulmonary metastases occurred in 39 (6.9%) patients. The smoking histories revealed 355 never smokers, 119 former smokers and 93 current smokers among the subjects. Multivariate analysis revealed that being a current smoker (hazard ratio = 2.72, 95% CI 1.18-6.25; P = 0.02) was an independent risk factor for pulmonary metastases. CONCLUSION: Smoking may be a risk factor for pulmonary metastasis of colorectal cancer. Cessation of smoking should be recommended to prevent pulmonary metastasis, although further basic and clinical studies are required.

Network meta-analysis of protocol-driven care and laparoscopic surgery for colorectal cancer.

BACKGROUND: Laparoscopic approaches and standardized recovery protocols have reduced morbidity following colorectal cancer surgery. As the optimal regimen remains inconclusive, a network meta-analysis was undertaken of treatments for the development of postoperative complications and mortality. METHODS: MEDLINE, Embase, trial registries and related reviews were searched for randomized trials comparing laparoscopic and open surgery within protocol-driven or conventional perioperative care for colorectal cancer resection, with complications as a defined endpoint. Relative odds ratios (ORs) for postoperative complications and mortality were estimated for aggregated data. RESULTS: Forty trials reporting on 11 516 randomized patients were included with the network. Open surgery within conventional perioperative care was the index for comparison. The OR relating to complications was 0·77 (95 per cent c.i. 0·65 to 0·91) for laparoscopic surgery within conventional care, 0·69 (0·48 to 0·99) for open surgery within protocol-driven care, and 0·43 (0·28 to 0·67) for laparoscopic surgery within protocol-driven care. Sensitivity analyses excluding trials of low rectal cancer and those with a high risk of bias did not affect the treatment estimates. Meta-analyses demonstrated that mortality risk was unaffected by perioperative strategy. CONCLUSION: Laparoscopic surgery combined with protocol-driven care reduces colorectal cancer surgery complications, but not mortality. The reduction in complications with protocol-driven care is greater for open surgery than for laparoscopic approaches. Registration number: CRD42015017850 (https://www.crd.york.ac.uk/PROSPERO).

Meta-analysis of the risk of small bowel obstruction following open or laparoscopic colorectal surgery.

BACKGROUND: One of the potential advantages of laparoscopic compared with open colorectal surgery is a reduction in postoperative bowel obstruction events. Early reports support this proposal, but accumulated evidence is lacking. METHODS: A systematic review and meta-analysis was performed of randomized clinical trials and observational studies by searching the PubMed and Cochrane Library databases from 1990 to August 2015. The primary outcomes were early and late postoperative bowel obstruction following laparoscopic and open colorectal surgery. Both ileus and bowel obstruction were defined as a postoperative bowel obstruction. Subgroup and sensitivity analyses were performed, and a random-effects model was used to account for the heterogeneity among the studies. RESULTS: Twenty-four randomized clinical trials and 88 observational studies were included in the meta-analysis; 106 studies reported early outcome and 12 late outcome. Collectively, these studies reported on the outcomes of 148 392 patients, of whom 58 133 had laparoscopic surgery and 90 259 open surgery. Compared with open surgery, laparoscopic surgery was associated with reduced rates of early (odds ratio 0·62, 95 per cent c.i. 0·54 to 0·72; P < 0·001) and late (odds ratio 0·61, 0·41 to 0·92; P = 0·019) postoperative bowel obstruction. Weighted mean values for early postoperative bowel obstruction were 8 (95 per cent c.i. 6 to 10) and 5 (3 to 7) per cent for open and laparoscopic surgery respectively, and for late bowel obstruction were 4 (2 to 6) and 3 (1 to 5) per cent respectively. CONCLUSION: The reduction in postoperative bowel obstruction demonstrates an advantage of laparoscopic surgery in patients with colorectal disease.

Adverse effects of long-term drain placement and the importance of direct aspiration: a retrospective cohort study.

BACKGROUND: Long-term placement of prophylactic drains may result in retrograde infections. AIM: To investigate the association between the timing of drain removal and clinical outcomes. METHODS: This retrospective, single-centre cohort study evaluated 110 patients who underwent elective gastrointestinal or hepatopancreatobiliary surgery and developed subsequent organ/space surgical site infection (SSI) between 2016 and 2020. The difference between the culture-positive species of prophylactic drains and direct aspiration was evaluated; whether the prophylactic drains functioned effectively at the time of SSI diagnosis; and whether the empirical antibiotics administered before drainage were effective against all the detected bacteria. Finally, clinical outcomes were compared between early (i.e. cases wherein the prophylactic drain had already been removed or replaced at the time of SSI diagnosis) and late (removal after diagnosis) drain removal. FINDINGS: The prophylactic drains functioned effectively in only 27 (25%) patients at the time of SSI diagnosis. Due to the results of direct aspiration cultures, 43% of patients required antibiotic escalation. The median time to drain removal or first replacement was seven postoperative days. The early removal group included 43 patients (39%). Compared with early removal, late removal resulted in a higher frequency of vancomycin use (7.0% vs 22.4%; P = 0.037). CONCLUSION: Prolonged prophylactic drain placement is associated with complicated infections requiring vancomycin; therefore, the drains should be removed as soon as possible. Additionally, obtaining the cultures of direct aspiration should be actively considered, as escalation of antimicrobial therapy is often performed based on culture results.

Safety and feasibility of laparoscopic surgery for elderly rectal cancer patients in Japan: a nationwide study.

BACKGROUND: This study aimed to analyse the perioperative results from a national dataset of rectal cancer resections in elderly patients. METHODS: The clinical records of patients undergoing rectal cancer surgery between 2012 and 2014 were retrieved from the Japanese National Clinical Database and analysed retrospectively. Patients were categorized according to age and those 80 years or older were defined as elderly. Subgroups were also defined according to the surgical approach (laparoscopy versus open surgery). The short-term outcomes, including mortality, anastomotic leak, surgical site infections and medical complications were compared between subgroups. RESULTS: Of 56 175 patients undergoing rectal cancer surgery, some 6717 patients were elderly and laparoscopy was performed in 46.8 per cent of the sample. When comparing laparoscopy and open surgery in elderly patients, the operative mortality rate (1.5 versus 2.8 per cent; P < 0.001), the incidence of anastomotic leakage (5.2 versus 6.5 per cent; P = 0.026), surgical site infections (6.0 versus 8.0 per cent; P = 0.001), pneumonia (1.4 versus 2.5 per cent; P = 0.001), renal failure (0.7 versus 1.3 per cent; P = 0.016) and cardiac events (0.3 versus 0.8 per cent; P = 0.008) were lower for laparoscopy than for open surgery. The overall complication rate in elderly patients (19.5 per cent) was comparable to that in the younger group (P = 0.07). However, incidence of systemic complications was significantly higher in elderly than in younger patients (all P < 0.001). CONCLUSION: Laparoscopy was safe and feasible in elderly patients compared with open surgery. However, the rates of systemic complications were significantly higher than in younger patients.

Effect of high ligation on survival of patients undergoing surgery for primary colorectal cancer and synchronous liver metastases.

BACKGROUND: Although R0 surgery is recommended for stage IV colorectal cancer, the degree of required lymphadenectomy has not been established. The aim of this study was to investigate the prognostic impact of high ligation (HL) of the feeding artery and the number of retrieved lymph nodes after R0 surgery for colorectal cancer and synchronous colorectal cancer liver metastasis (CRLM). METHODS: This was a multi-institutional retrospective analysis of patients with colorectal cancer and synchronous CRLM who had R0 surgery between January 1997 and December 2007. Clinical and pathological features were compared in patients who underwent HL and those who had a low ligation (LL). Kaplan-Meier analysis was performed to estimate the effect of HL on overall survival (OS). The impact of several risk factors on survival was analysed using the Cox proportional hazards model. RESULTS: Of 549 patients, 409 (74·5 per cent) had HL. Median follow-up was 51·4 months. HL significantly improved the 5-year OS rate (58·2 per cent versus 49·3 per cent for LL; P = 0·017). Multivariable analysis revealed HL to be a significant prognostic factor compared with LL (5-year mortality: hazard ratio (HR) 0·68, 95 per cent c.i. 0·51 to 0·90; P = 0·007). In subgroup analysis, the positive effect of HL on OS was greatest in patients with lymph node metastasis. CONCLUSION: HL of the feeding artery was associated with improved OS in patients with colorectal cancer and synchronous CRLM after R0 surgery.

ANTECEDENTES: Aunque se recomienda una cirugía R0 para el cáncer colorrectal (colorectal cancer, CRC) en estadio IV, no se ha establecido el grado de linfadenectomía requerida. El objetivo de este estudio fue investigar el impacto pronóstico de la ligadura alta (high ligation, HL) de la arteria que irriga el tumor y el número de ganglios linfáticos (lymph nodes, LN) identificados después de cirugía R0 en pacientes con cáncer colorrectal y metástasis hepáticas sincrónicas (colorectal cancer liver metastasis, CRLM). MÉTODOS: En este estudio se realizó un análisis retrospectivo multicéntrico de pacientes con CRC y CRLM sincrónicas en los que se realizó una cirugía R0 desde enero de 1997 hasta diciembre de 2007. Se compararon las características clínicas y patológicas entre los pacientes a los que, durante la cirugía R0, se practicó una HL frente a los que no se practicó esta técnica. El análisis de Kaplan-Meier se realizó para estimar el efecto de la HL en la supervivencia global (overall survival, OS). El impacto de varios factores de riesgo sobre la supervivencia se analizó utilizando el modelo de Cox de riesgo proporcional. RESULTADOS: Sobre un total de 549 pacientes, se realizó una HL en 409 (74,5%), y el período de seguimiento medio en esta cohorte fue de 51,4 meses. La HL mejoró significativamente la tasa de OS a los 5 años (HL 37,7% versus no HL 27,1%, P = 0,02). El análisis multivariable mostró que la HL era un factor pronóstico significativo en comparación con la no realización de una HL (cociente de riesgos instantáneos, hazard ratio, HR de muerte a 5 años = 0,68 (i.c. del 95% 0,51-0,90), P < 0,01)). En el análisis de subgrupos, el efecto positivo de la HL sobre la OS fue mayor en pacientes con metástasis ganglionares. CONCLUSIÓN: La ligadura alta de la arteria que irriga el tumor se asoció con una mejor OS en pacientes con CRC y CRLM sincrónicas después de una cirugía R0.

A case of laparoscopic surgery for blue rubber bleb nevus syndrome.

A 33-year-old woman had presented with some cutaneous and gastrointestinal hemangiomas with iron deficiency anemia three years previously and blue rubber bleb nevus syndrome was diagnosed. Although she had taken an iron supplement for three years, she hoped to discontinue it and underwent laparoscopic surgery for nine gastrointestinal hemangiomas. She was discharged on the ninth postoperative day without any complications. She has been fit without iron deficiency anemia for a year since the operation. Blue rubber bleb nevus syndrome is a rare disease accompanied with cutaneous and gastrointestinal vascular malformations and about 150 cases have been reported. To the best of our knowledge, this is the first report of blue rubber bleb nevus syndrome with small and large intestinal lesions that were treated with laparoscopic surgery.

Covalent modification of all members of human cullin family proteins by NEDD8.

Recently we found that NEDD8, a ubiquitin-like protein, was linked covalently to human cullin-4A (abbreviated Cul-4A) by a new ubiquitin-related pathway that is analogous to but distinct from the ligating system for SUMO1, another ubiquitin-like protein. However, it remained unknown whether the other five members of the family of human cullin/Cdc53 proteins are modified by NEDD8. Here we report that all Hs-Cul family proteins, such as Cul-1, Cul-2, Cul-3, Cul-4B, and Cul-5, in addition to Cul-4A, were modified by covalent attachment of NEDD8 in rabbit reticulocyte lysates. Moreover, by comprehensive Northern-blot analyses, we examined multiple tissue distributions of the messages for all Cul-family proteins, NEDD8, and the NEDD8-ligating system consisting of APP-BP1/hUba3, and hUbc12, which function as E1- and E2-like enzymes, respectively. The expressions of Cul-1, Cul-2, and Cul-3 resembled each other and were apparently correlated to those of NEDD8 and the NEDD8-ligating system in various human cells and tissues. However, the mRNA levels of Cul-4A, Cul-4B, and Cul-5 differed considerably from each other as well as from other Cul-family proteins. The enhanced expression of all Cul-family proteins except Cul-5 was observed in a variety of tumor cell lines.

Comparison of oncological safety between nipple sparing mastectomy and total mastectomy using propensity score matching.

INTRODCUTION: Although nipple sparing mastectomy (NSM) has attracted increased recognition as an alternative to traditional mastectomy approaches, its oncological safety is unclear. The purpose of this study was to compare the local recurrence rate between NSM and total mastectomy (TM). METHODS: Between 2003 and 2013, 121 and 557 patients with stage 0-III breast cancer underwent NSM and TM respectively. Multivariate Cox regression and propensity score models were used to compare the two groups. RESULTS: There was no significant difference in the five-year local recurrence rate between the NSM and TM groups (7.6% vs 4.9%, p=0.398). In multivariate analysis, NSM was not a risk factor for local recurrence (hazard ratio: 1.653, 95% confidence interval: 0.586-4.663, p=0.343). Propensity score matching found similar five-year local recurrence free survival rates between the two groups (92.3% vs 93.7%, p=0.655). CONCLUSIONS: Our results suggest that NSM may provide oncological safety comparable with mastectomy for carefully selected patients.

Secretory production of recombinant urokinase-type plasminogen activator-annexin V chimeras in Pichia pastoris.

To produce a thrombi-targeting plasminogen activator, we expressed a fused gene that contains a modified pre-sequence of Mucor pussilus rennin (MPR) followed by a chimeric gene of single-chain urokinase-type plasminogen activator (scu-PA)::annexin V (AV). The fused gene was ligated into an integrative vector, under the control of the alcohol oxidase 1 (AOX1) promoter (p), and transformed into Pichia pastoris. Transformants were monitored for the secretion of fibrinolytic activity. The highest expressing clone, HB225, secreted as much as 600 international units (IU) of fibrinolytic activity per ml of culture medium under optimal conditions. It contained three tandem copies of the full-size vector disruptively integrated into the AOX1 sequence. Western blot analysis revealed that the secreted chimera was highly susceptible to proteolysis. Addition of excess amino acids (aa) to the culture medium minimized the degree of proteolysis. Two major species of chimera, 85 and 65 kDa, were then isolated from the culture medium. The former was the intact form consisting of a single-chain and showing full enzyme activity after activation by plasmin. The latter was an enzymatically processed form consisting of two chains held by a disulfide bond, having full enzyme activity without activation. Both chimeras exhibited calcium-dependent phospholipid (PL)-binding affinities similar to the parent AV.

mRNA expression and protein localization of dentin matrix protein 1 during dental root formation.

Dentin matrix protein 1 (DMP1) is an acidic phosphoprotein. DMP1 was initially detected in dentin and later in other mineralized tissues including cementum and bone, but the DMP1 expression pattern in tooth is still controversial. To determine the precise localization of DMP1 messenger RNA (mRNA) and the protein in the tooth, we performed in situ hybridization and immunohistochemical analyses using rat molars and incisors during various stages of root formation. During root dentin formation of molars, DMP1 mRNA was detected in root odontoblasts in parallel with mineralization of the dentin. However, the level of DMP1 mRNA expression in root odontoblasts decreased near the coronal part and was absent in coronal odontoblasts. DMP1 protein was localized along dentinal tubules and their branches in mineralized root dentin, and the distribution of DMP1 shifted from the end of dentinal tubules to the base of the tubules as dentin formation progressed. During the formation of the acellular cementum, DMP1 mRNA was detected in cementoblasts lining the acellular cementum where its protein was localized. During the formation of the cellular cementum, DMP1 mRNA was detected in cementocytes embedded in the cellular cementum but not in cementoblasts, and its protein was localized in the pericellular cementum of cementocytes including their processes. During dentin formation of incisors, DMP1 mRNA was detected in odontoblasts on the cementum-related dentin, where its protein was localized along dentinal tubules near the mineralization front. The localization of DMP1 mRNA and protein in dentin and cementum was related to their mineralization, suggesting that one of the functions of DMP1 may be involved in the mineralization of dentin and cementum during root formation.

Attenuation of rat lung isograft reperfusion injury with a combination of anti-ICAM-1 and anti-beta2 integrin monoclonal antibodies.

Four different combinations of monoclonal antibodies against rat ICAM-1, CD-11a, and CD-18 were utilized to determine the relative importance of LFA-1, Mac-1, and ICAM-1 in a rat model of severe lung allograft reperfusion injury. Negative control animals were given phosphate buffered saline (the carrier solution for the antibodies), while positive control animals were rendered neutropenic by the administration of a polyclonal mouse IgG. Antibodies were given with the donor lung flush, prior to left lung graft reperfusion, or both. Isolated graft function was determined 24 hr after implantation by arterial blood gas (ABG), and after sacrifice the native and transplanted lungs underwent bronchoalveolar lavage for alveolar protein quantitation, cell count and differential, and myeloperoxidase assay. Additionally, whole lung homogenates were assayed for myeloperoxidase activity. We found that the combination of anti-ICAM-1 (1 mg/kg) added to the donor lung flush, and anti-CD11a, anti-CD18, and anti-ICAM-1 (2 mg/kg i.v. of each) given to the recipient prior to reperfusion, resulted in significantly improved lung graft pAO2 by ABG, and decreased alveolar protein, cell count, and myeloperoxidase activity compared with control animals. Improvement was less than that seen in the neutropenic recipients, however. We conclude that LFA-1, Mac-1, and ICAM-1 are all important adhesion molecules in lung allograft reperfusion injury--yet even with antibody blockade of all three there are additional mechanisms allowing for neutrophil influx into the lungs.

Hyperinflation of canine lung allografts during storage increases reperfusion pulmonary edema.

The optimal state of inflation for lung allografts during preservation is not known. We previously showed that hyperinflation of canine lung allografts during storage improved posttransplant graft function as measured during 10 minutes of contralateral pulmonary artery occlusion. However, we have also shown that hyperinflation during storage increases pulmonary capillary permeability. It is possible that short-term total cardiac perfusion through the transplanted left lung (for assessment) may not adequately reproduce the clinical situation. The purpose of this study was to assess the effects of hyperinflation during storage in a canine left single-lung transplantation model in which all perfusion was continuously directed to the graft after implantation. Twenty canine left single-lung transplants were done. The lungs of donor animals were ventilated at a tidal volume of 750 ml and an inspired oxygen fraction of 100%. Donor lungs were flushed with modified Euro-Collins solution and the trachea occluded at end inspiration. For donors in groups I and III, the trachea was sealed at that postinflation volume. In groups II and IV, 200 cc of air was withdrawn from the endotracheal tube under positive pressure and the trachea sealed at the lower tidal volume. Lungs were then extracted and stored at 1 degree C for 12 hours. After the preservation period, left lung transplants were performed. After implantation in all groups, the right pulmonary artery was ligated. In groups I and II, the right bronchus was ligated and in groups III and IV the right bronchus was kept open. Subsequent allograft gas exchange and hemodynamics were assessed during a 6-hour period of reperfusion. After assessment, both lungs were excised, wet/dry lung weight ratio was calculated, and histologic examination was done. During the 6-hour assessment, lungs stored in a state of hyperinflation (groups I and III) showed rapid deterioration of gas exchange. At the final assessment, arterial oxygen tension and alveolar-arterial oxygen gradient of groups I and III were significantly worse than those of groups II and IV (group I versus group II: arterial oxygen tension 87.5 +/- 15.0 versus 373.8 +/- 65.5 mm Hg, alveolar-arterial oxygen gradient 564.4 +/- 13.2 versus 298.6 +/- 69.3 mm Hg, p < 0.05; group III versus group IV: arterial oxygen tension 245.4 +/- 33.0 versus 543.6 +/- 41.8 mm Hg, alveolar-arterial oxygen gradient 392.5 +/- 35.6 versus 120.5 +/- 34.7 mm Hg, p < 0.01). We conclude that donor lung hyperinflation during storage does not provide better posttransplant allograft function when perfusion is limited only to the allograft.

Experimental carinal autotransplantation and allotransplantation.

Grafting is required when a primary reconstruction of a carinal defect is not feasible. A series of experiments in 21 dogs was conducted to assess the possibility of carinal reconstruction with the use of carinal autografts and allografts, with or without omentopexy. Carinal autograft transplantations were done without omentopexy in group A (n = 6) and with omentopexy in group B (n = 6). In group C (n = 9), carinal allograft transplantations were done with omentopexy and FK 506 was administered after operation. Survival of grafts was seen in 50% of group A, 83% of group B, and 44% of group C dogs. Postoperative bronchoscopy revealed inflammatory changes in the surviving grafts; the changes resolved more rapidly in the dogs with omentopexy than in the dogs without omentopexy. These experiments suggest that omentopexy is an effective method of facilitating survival and healing in carinal grafts and that carinal reconstruction with carinal allografts with FK 506 is feasible.

Tracheal transplantation for carinal reconstruction in dogs.

BACKGROUND: Experimental carinal allotransplantation has been performed with tracheocarinal Y-shaped allografts in dogs. In this study we tried canine carinal reconstruction with cylindrical allografts. MATERIAL AND METHODS: Carinal reconstruction was performed with allotransplantation of cylindrical trachea in dogs, and graft healing was evaluated by bronchoscopic observation, mucosal blood flow measurement, and histologic examination. A section of the recipient carina containing five tracheal rings and two main stem bronchi was removed, and a donor trachea seven rings long was inserted between the recipient trachea and the left main stem bronchus; then side-to-end anastomosis was performed between the graft midportion and recipient right main stem bronchus (new carina). The grafts were wrapped with pedicled omentum. Fresh grafts were transplanted into one group of dogs (n=8 ), and grafts cryopreserved for 1 week were transplanted into another group (n=7). RESULTS: No anastomotic leakage occurred in any dog. Excellent healing of grafts and graft anastomoses was observed by fiberoptic bronchoscopy in six dogs (75%) in the fresh graft group and in four dogs (57%) in the cryopreserved graft group. The mucosal blood flow in the new carina decreased remarkably and, although it recovered, mucosal blood flow remained under the preoperative level on day 28 after the operation. CONCLUSION: Cylindrical tracheal allotransplantation is useful for carinal reconstruction, and the method of side-to-end anastomosis between the donor trachea and recipient bronchus is a feasible and accessible procedure in dogs.

Inhaled nitric oxide improves lung allograft function after prolonged storage.

Morbidity caused by early allograft dysfunction, manifested by a progressive increase in pulmonary vascular resistance and a decrease in oxygenation, remains a serious problem in lung transplantation. Inhalation of nitric oxide, an essential homeostatic molecule, has been shown to have beneficial effects on a variety of acute lung injuries. The purpose of the present study was to investigate the effect of inhaled nitric oxide on posttransplant function of canine left lung allografts. Fourteen dogs underwent left lung allotransplantation. Donors received systemic heparin and prostaglandin E1 followed by pulmonary artery flush with modified Euro-Collins solution. Donor left lungs were stored for 18 hours at 1 degree C and subsequently implanted. Immediately after reperfusion, the contralateral right main pulmonary artery and bronchus were ligated. The chest was closed and recipients turned to the supine position for the 6-hour assessment period. Hemodynamic and arterial and venous blood gas analyses were made at 15-minute intervals at an inspired oxygen fraction of 1.0 and 5 cm of water positive end-expiratory pressure. Animals were killed at the end of the assessment. Allograft myeloperoxidase activity assays and wet/dry weight ratios were done. In group I (n = 5), nitric oxide gas was administered continuously at concentrations of 60 to 70 ppm before reperfusion and throughout the 6-hour assessment period. In group II (n = 5), nitric oxide administration was initiated at the same concentration after reperfusion injury had developed. Group III animals (n = 4) received no nitric oxide. Significant improvement in gas exchange was apparent in group I. At the end of the 6-hour assessment period, mean arterial oxygen tension was 253.8 +/- 44.7 mm Hg and 114.9 +/- 25.5 mm Hg in groups I and III, respectively (p < 0.05). Group II animals had no improvement in oxygenation with nitric oxide. Systemic hemodynamics were unaffected by nitric oxide. However, an immediate decrease in pulmonary vascular resistance was noted. Group I myeloperoxidase activity was significantly lower than that in control group III (0.24 +/- 0.06 versus 0.36 +/- 0.04 units, respectively; p < 0.05).

Inhaled nitric oxide does not increase rat pulmonary allograft rejection.

BACKGROUND: There is evidence that inducible nitric oxide (NO) may be directly related to the process of allograft rejection. Because of its strong pulmonary vasodilatory activity, inhaled NO (INO) has recently been used as a therapeutic option for allograft dysfunction after lung transplantation. The action of inducible NO and inhaled NO seems contradictory for preserving posttransplantation pulmonary allograft function. INO used for lung transplant recipients may actually enhance acute allograft rejection. We studied the effect of INO on acute allograft rejection with a rat pulmonary allograft model. METHOD: A total of 24 left lung allotransplantations were performed from Lewis donors into F344 recipients. Animals were divided into two groups and inhaled either room air alone or 20 ppm NO with room air in a closed chamber immediately after transplantation until rats were killed on days 7 and 14. During observation, NO uptake was monitored by measuring serum NO2-/NO3- level. Acute rejection was evaluated by use of a semiquantitative radiographic scoring method (aeration score: 0 to 6, opaque to normal appearance) and rejection score (0 to 4, no sign of rejection to diffuse mononuclear infiltration). RESULTS: Markedly elevated serum NO2-/NO3- levels were observed in the NO inhalation group compared with levels in the normal air inhalation control group (110.8 +/- 25.3 vs 16.3 +/- 4.0 micromol/L/ml on day 7, p < 0.01; 107.0 +/- 30.9 vs 16.8 +/- 4.8 micromol/L/ml on day 14, p < 0.01). However, no positive effect of INO on acute rejection was found histologically or radiographically. CONCLUSION: The effect of INO on acute rejection is likely so minimal as not to be clinically relevant.

Partial purification and properties of citrate synthase from sea urchin eggs.

Citrate synthase [EC 4.1.3.7] was purified from sea urchin eggs about 14-fold with a 23% yield, based on the activity of the crude extract. The molecular weight of the enzyme was about 100,000 as determined by gel filtration. The optimum pH was about 7.8 in 100 mM Tris-HCl. The apparent Km values for acetyl-CoA and for oxaloacetate were 33 and 3.2 muM, respectively. Monovalent and divalent cations inhibited the enzyme. Iodoacetamide, pCMB, EDTA, NaF, and dithiothreitol did not affect the enzyme activity. Oxaloacetate protected the enzyme against heat denaturation. Among nucleotides, ATP was the most potent inhibitor of the enzyme. The inhibition by ATP was competitive with respect to acetyl-CoA and mixed with respect to oxaloacetate.

Purification and properties of mitochondrial malate dehydrogenase from unfertilized eggs of the sea urchin, Anthocidaris crassispina.

Purification and characterization of mitochondrial malate dehydrogenase [EC 1.1.1.37] from unfertilized eggs of the sea urchin, Anthocidaris crassispina, are described. The purification method consisted of dextran sulfate fractionation, Blue Dextran Sepharose chromatography, Phenyl-Sepharose hydrophobic chromatography and DEAE-cellulose chromatography. The enzyme was purified 771-fold with a 7% yield from the crude extract. The purified enzyme appeared homogeneous on polyacrylamide gel electrophoresis under both native and denatured conditions. After incubation at 45 degrees C for 50 min, the enzyme lost about 90% of its activity. In the presence of NADH, however, the enzyme was protected against the heat denaturation. The native enzyme had a molecular weight of about 65,000 and probably consisted of two identical subunits. In the reduction of oxaloacetate with NADH, a broad optimum pH ranging from 8.2 to 9.4 was found with 50 mM Tris-HCl and glycine-NaOH buffers. Sodium phosphate buffer apparently activated the enzyme. The apparent Km values for oxaloacetate and NADH were 19 microM and 30 microM, respectively. The optimum pH for malate oxidation with NAD+ was 10.2 in 50 mM NaHCO3-Na2CO3 buffer. The apparent Km values for malate and NAD+ were 7.0 mM and 0.6 mM, respectively. Zinc ion, sulfite ion, p-chloromercuriphenylsulfonate and adenine nucleotides strongly inhibited the enzyme.