An estimate of hernia prevalence in Sierra Leone from a nationwide community survey.

PURPOSE: A large number of unrepaired inguinal hernias is expected in sub-Saharan Africa where late presentation often results in incarceration, strangulation, or giant scrotal hernias. However, no representative population-based data are available to quantify the prevalence of hernias. We present data on groin masses in Sierra Leone to estimate prevalence, barriers to care, and associated disability. METHODS: A cluster randomized, cross-sectional household survey of 75 clusters of 25 households with 2 respondents each was designed to calculate the prevalence of and disability caused by groin hernias in Sierra Leone using a verbal head-to-toe examination. Barriers to hernia repairs were assessed by asking participants the main reason for delay in surgical care. RESULTS: Information was obtained from 3,645 respondents in 1,843 households, of which 1,669 (46%) were male and included in the study. In total, 117 males or 7.01% (95% CI 5.64-8.38) reported a soft or reducible swelling likely representing a hernia with four men having two masses. Of the 93.2% who indicated the need for health care, only 22.2% underwent a procedure, citing limited funds (59.0%) as the major barrier to care. On disability assessment, 20.2% were not able to work secondary to the groin swelling. CONCLUSIONS: The results indicate groin masses represent a major burden for the male population in Sierra Leone. Improving access to surgical care for adult patients with hernias and early intervention for children will be vital to address the burden of disease and prevent complications or limitations of daily activity.

Care within a veterans hospital: earlier detection of colon cancer.

BACKGROUND: In 1998 the Veterans Administration mandated an externally monitored targeted colon cancer screening rate that was expected to result in earlier cancer detection and improved patient survival. The effectiveness of the protocol was evaluated in a retrospective case series at a tertiary care Veterans Administration Hospital that included all patients with the diagnosis of colon cancer between 1991 and 2003. METHODS: Tumor stage, tumor location, and patient survival data were recorded and compared to National Cancer Data Base (NCDB) benchmarks. RESULTS: The study facility had a greater percentage of early cancers and fewer later stage cancers than the NCDB benchmark. Overall survival was better for the VA cohort compared to NCDB (all-cause 5-year survival: VA, 0.72; NCDB, 0.47. p < or = .001). CONCLUSIONS: The VA facility had a significantly greater percentage of early cancers and fewer stage III or IV cancers compared to a national benchmark and significantly improved survival compared to the national benchmark.

Transnasal small-caliber esophagogastroduodenoscopy for preoperative evaluation of the high-risk morbidly obese patient.

BACKGROUND: Esophagogastroduodenoscopy (EGD) is an important facet of the preoperative evaluation for bariatric surgery. Morbidly obese patients are at high risk for airway complications during this procedure, and an attractive alternative is transnasal EGD. This report describes a series of patients evaluated successfully using this technique. METHODS: All patients undergoing preoperative transnasal small-caliber EGD for morbid obesity surgery between September 2004 and June 2005 at a Veterans Affairs Hospital were included in the analysis. The variables assessed were the adequacy of the examination, patient tolerance, the need for sedation, and the ability to perform interventions. RESULTS: The study enrolled 25 patients (17 men and 8 women) with an average age of 55 years (range, 44-63 years) and an average body mass index (BMI) of 47 kg/m2 (range, 38-69 kg/m2). All the patients met the 1991 National Institutes of Health (NIH) Consensus Conference Criteria for bariatric surgery and were undergoing preoperative evaluation. The most common comorbidities were hypertension (82%), diabetes mellitus (80%), and obstructive sleep apnea (68%). All 25 patients had successful cannulation of the duodenum's second portion with excellent tolerance. There were no sedation requirements for 23 (92%) of the 25 patients. Significant pathology was found in 14 (56%) of the 25 patients, including hiatal hernia (28%), gastritis (16%), esophageal intestinal metaplasia (16%), esophagitis (12%), gastric polyps (8%), gastric ulcer (4%) and esophageal varices (4%). Biopsies were indicated for 12 patients and successful for all 12 (100%). CONCLUSION: Transnasal small-caliber EGD is a feasible and safe alternative to conventional EGD for the preoperative evaluation of patients undergoing bariatric surgery. It requires minimal to no sedation in a population at high risk for complications in this setting. In addition, this technique is effective in identifying pathology that requires preoperative treatment and offers a complete examination with biopsy capabilities. This technique should be considered for all morbidly obese patients at high risk for airway compromise during EGD.

Cytomegalovirus Retinitis in infancy.

PURPOSE: To describe the presentation of cytomegalovirus retinitis (CMVR) in a series of infants. METHODS: Immunocompromised infants with either HIV or systemic cytomegalovirus (CMV) were examined for CMVR. Ocular involvement was recorded and monitored by digital imaging. RESULTS: Five infants were detected to have CMVR. All the infants demonstrated changes within the macula. One infant progressed from a fine granular pattern to fulminant CMVR. CONCLUSION: Infants under a year with CMVR have a predilection for the disease to present at the macula, in contrast to the presentation in adults, which tends to involve more peripheral parts of the retina.

Decompressive colonoscopy with intracolonic vancomycin administration for the treatment of severe pseudomembranous colitis.

BACKGROUND: We explored the potential of early decompressive colonoscopy with intracolonic vancomycin administration as an adjunctive therapy for severe pseudomembranous Clostridium difficile colitis with ileus and toxic megacolon. METHODS: We reviewed the symptoms, signs, laboratory tests, radiographic findings, and outcomes from the medical records of seven patients who experienced eight episodes of severe pseudomembranous colitis with ileus and toxic megacolon. All seven patients underwent decompressive colonoscopy with intracolonic perfusion of vancomycin. RESULTS: Fever, abdominal pain, diarrhea, abdominal distention, and tenderness were present in all patients. Five of seven patients were comatose, obtunded, or confused, and six of the seven required ventilatory support. The white blood cell count was greater than 16,000 in seven cases (six patients). Colonoscopy showed left-side pseudomembranous colitis in one patient, right-side colitis in one patient, and diffuse pseudomembranous pancolitis in five patients. Two patients were discharged with improvement. Five patients had numerous medical problems leading to their death. Complete resolution of pseudomembranous colitis occurred in four patients. One patient had a partial response, and two patients failed therapy. CONCLUSION: Colonoscopic decompression and intracolonic vancomycin administration in the management of severe, acute, pseudomembranous colitis associated with ileus and toxic megacolon is feasible, safe, and effective in approximately 57% to 71% of cases.

Results of a screening program for prostate cancer in patients scheduled for abdominoperineal resection for colorectal pathologic findings.

OBJECTIVES: Because of the difficulty of accessing the prostate for tissue sampling after surgical removal of the rectum and obliteration of the anus, we started an early detection program for prostate cancer in all men scheduled for abdominoperineal resection. METHODS: Twenty consecutive men were screened for prostatic adenocarcinoma before planned abdominoperineal resection for colorectal pathologic findings. Patients were 48 to 77 years old (mean 66.9). Screening included serum prostate-specific antigen determination and digital rectal examination. Those patients with suspicious findings underwent transrectal ultrasound-guided sextant biopsies of the prostate. RESULTS: One patient was excluded because of a prior history of prostate cancer. Six (31.6%) of the remaining 19 patients demonstrated elevated prostate-specific antigen levels (greater than 4.0 ng/mL); two of these patients also had an abnormal digital rectal examination. Transrectal ultrasound and prostate biopsies in these 6 patients revealed prostatic adenocarcinoma in 3 patients (50% of those undergoing biopsies or 15.8% of those screened). The 13 patients who did not undergo prostate biopsies had prostate-specific antigen levels from 0.4 to 2.4 ng/mL (mean 0.9) and normal prostate glands according to the digital rectal examinations. CONCLUSIONS: Screening for prostate cancer in men 50 years old or older with 10 years or longer life expectancy before they undergo abdominoperineal resection detects a significant number of prostatic malignancies and should be encouraged.

External beam radiotherapy for synchronous rectal and prostatic tumors.

Two patients were diagnosed with large rectal tumors and localized prostate cancer. The prostate-specific antigen level at diagnosis was 7.9 ng/mL and 9.0 ng/mL in the 2 patients. Knowledge of the presence of both tumors and their close proximity allowed creation of a modified three-dimensional conformal radiotherapy plan to treat both tumors. The patients had no evidence of rectal tumor recurrence and their prostate-specific antigen level was 0.5 ng/mL and 0.7 ng/mL at 1 and 2 years after therapy, respectively. We conclude that efficient, effective pelvic irradiation can be designed for synchronous rectal and prostate malignancies.

Does an infected peripancreatic fluid collection or abscess mandate operation?

OBJECTIVE: To assess the treatment of peripancreatic fluid collections or abscess with percutaneous catheter drainage (PCD). SUMMARY BACKGROUND DATA: Surgical intervention has been the mainstay of treatment for infected peripancreatic fluid collections and abscesses. Increasingly, PCD has been used, with mixed results reported in the literature. METHODS: A retrospective chart review of 1993 to 1997 was performed on 82 patients at a tertiary care public teaching hospital who had computed tomography-guided aspiration for suspected infected pancreatic fluid collection or abscess. Culture results, need for subsequent surgical intervention, length of stay, and death rate were assessed. RESULTS: One hundred thirty-five aspirations were performed in 82 patients (57 male patients, 25 female patients) with a mean age of 40 years (range 17-68). The etiologies were alcohol (41), gallstones (32), and other (9). The mean number of Ranson's criteria was four (range 0-9). All patients received antibiotics. Forty-eight patients had evidence of pancreatic necrosis on computed tomography scan. Cultures were negative in 40 patients and positive in 42. Twenty-five of the 42 culture-positive patients had PCD as primary therapy, and 6 required subsequent surgery. Eleven patients had primary surgical therapy, and five required subsequent surgery. Six patients were treated with only antibiotics. The death rates were 12% for culture-positive patients and 8% for the entire 82 patients. CONCLUSIONS: Historically, patients with positive peripancreatic aspirate culture have required operation. This series reports an evolving strategy of reliance on catheter drainage. PCD should be considered as the initial therapy for culture-positive patients, with surgical intervention reserved for patients in whom treatment fails.

Antiviral cytotoxic activity across a species barrier in mixed xenogeneic chimeras: functional restriction to host MHC.

Reconstitution of lethally irradiated mice with a mixture of mouse and rat bone marrow cells (mouse + rat-->mouse) results in mixed xenogeneic chimerism and donor-specific tolerance. The current study demonstrates that mouse and rat T lymphocytes that have developed in xenogeneic chimeras are restricted to Ag presentation by mouse, but not rat, APC. Restriction to host Ags results in functional immunocompetence with generation of antiviral cytotoxic activity in vivo, within and across species barriers. These data demonstrate for the first time that the host thymus is sufficient to support development and positive selection of functional cross-species T lymphocytes. The superior immunocompetence, as compared with fully xenogeneic (rat-->mouse) chimeras, may prove to be of significant benefit in the clinical application of xenotransplantation to solid organ transplantation and immune reconstitution for AIDS.

Is cryosurgical ablation appropriate for treating hepatocellular cancer?

OBJECTIVE: To examine the feasibility and efficacy of cryosurgical ablation as treatment for patients with cirrhosis with unresectable hepatocellular carcinoma. DESIGN: Retrospective case series. SETTING: A tertiary public hospital and a cancer center. PATIENTS: Twelve patients with cirrhosis with hepatocellular carcinoma (stage II, 2; stage III, 1; stage IVA, 7; stage IVB, 2). INTERVENTIONS: Cryosurgical ablation of all identifiable tumors. Nine patients treated with curative intent were included in the survival analysis, and 3 were treated for palliation. Five patients were treated with preoperative intra-arterial chemoembolization. MAIN OUTCOME MEASURES: Perioperative complications and the effects of tumor stage and chemoembolization were examined. Patient survival and disease-free interval were calculated by life-table analysis. RESULTS: No perioperative deaths occurred and 1 patient had 2 postoperative complications: pneumonia and biloma. The mean survival has been 19 months after cryosurgical ablation and 29 months after diagnosis. Three of the 9 patients treated with curative intent died with recurrence at a mean of 17 months after cryosurgical ablation. Four patients are alive with recurrence at a mean of 19 months after cryosurgical ablation and 38 months after diagnosis. Two patients with stage II disease have no evidence of recurrence 10 and 32 months after cryosurgical ablation. CONCLUSIONS: Cryosurgical ablation is feasible and safe for treatment of hepatocellular carcinoma in patients with cirrhosis. The technique is primarily palliative but may provide a possibility of cure in patients with lower-stage disease.

Color flow sonography in evaluating the resectability of periampullary and pancreatic tumors.

Over the past several years, we have developed a technique to assess the resectability of periampullary and pancreatic tumors using color flow sonography. This is a feasibility study to determine if sonography with color flow imaging can play a role in evaluating patients with periampullary and pancreatic tumors. This study comprises a retrospective analysis of 51 patients referred for color flow sonographic evaluation of resectability of periampullary and pancreatic neoplasm. Scanning was performed with state-of-the-art color flow sonographic systems. Vessels that were touched or occluded by tumor were categorized according to a Pancreatic Color Doppler Score. Other factors affecting resectability (metastasis, enlarged nodes) were recorded. Sonographic findings were correlated with surgical resectability and pathologic findings regarding tumor margins. The color flow study was complete technically (all index vessels visualized) in 49 of 51 patients (96%). In all, 643 of 647 vessels (99.4%) were imaged. Forty-five patients had sufficient surgical, pathologic, or clinical proof to be included in the resectability analysis. All 18 patients with circumferential tumor or vascular occlusion (Pancreatic Color Doppler Score 4 and 5) were found to have unresectable disease. All 10 patients in whom tumor did not touch (Pancreatic Color Doppler Score 0) had negative margins. All 30 patients considered to have unresectable tumors sonographically could not be resected. Patients in this category had one or more of the following: positive pathologic margins, positive nodes, liver metastasis, or not clinically resectable. Six of 15 considered resectable sonographically (40%) were unresectable for cure. Surgeons believed that color flow sonography influenced management in 10 of 45 patients (22% overall). Color flow sonography, a painless, noninvasive, and relatively inexpensive examination, may be an effective screening tool to evaluate periampullary and pancreatic neoplasms for resectability. Our data show that color flow sonography can correctly predict unresectability of periampullary and pancreatic neoplasms. Any future evaluation of imaging and management of periampullary and pancreatic tumors should include color flow sonography.

Selective management of hepatic adenomas.

Hepatic adenomas are uncommon hepatic neoplasms that may be identified after life-threatening hemorrhage, or as an incidental radiologic finding. The incidence of malignant transformation is unknown, and the correct treatment strategy is unclear. We examined our 10-year experience in the management of 12 patients with hepatic adenomas. Eleven adults (mean age of 37.6 years) and one 3-month-old were identified. Nine of 10 adult females (90%) were taking a hormonal preparation at the time of diagnosis. Four patients with tumor sizes of 1.0 to 4.0 cm were observed after cessation of oral contraceptives. Four patients with lesions of 5.5 to 13 cm underwent surgical resection. Three had malignant transformation, and two of the three had increased Alpha-fetoprotein levels. Four patients presented with acute hemorrhage and were treated initially by hepatic arterial embolization. We conclude that management of adenomas should be individualized based on their size and mode of presentation. Patients with lesions less than 5 cm and normal alpha-fetoprotein can be safely observed off oral contraceptives and followed by radiologic imaging. Lesions >5 cm should be considered for surgical resection due to the risk of malignancy. Hepatic arterial embolization is a new approach for acute hemorrhage.

Assessment of resectability of pancreatic head and periampullary tumors by color flow Doppler sonography.

OBJECTIVE: To examine the sensitivity of color flow Doppler ultrasonography in assessing resectability of pancreatic head and periampullary tumors. DESIGN: Validation cohort study. SETTING: Tertiary care public hospital. PATIENTS: Thirty-seven patients with pancreatic head or periampullary cancer were studied by color flow Doppler examination of the relevant blood vessels. MAIN OUTCOME MEASURE: A pancreatic Doppler score (PDS) was defined as the closest circumferential contact of the tumor to the superior mesenteric vein, superior mesenteric artery, or portal vein. A PDS of 1 indicated no contact (n = 9); PDS 2, less than 50% contact (n = 10); PDS 3, 50% to 99% contact (n = 7); and PDS 4, encasement (n = 11). The PDS was compared with operative and histologic resection margins. RESULTS: The lack of vascular invasion was confirmed operatively in 7 of 7 patients with a PDS of 1, and 6 patients who underwent resection had clear histologic margins. Nine (90%) of 10 patients with a PDS of 2 were confirmed to have no vascular invasion, and 3 (43%) of 7 patients who underwent resection had clear margins. Five (83%) of 6 patients with a PDS of 3 had correct operative findings, and both patients who underwent resection had positive margins. Operative confirmation of encasement was found in all 7 patients with a PDS of 4 who had operative exploration, and none underwent resection. CONCLUSIONS: Color flow Doppler sonography and PDS predicted resectability and the histologic margin status (positive predictive value, 97%). Patients with a PDS of 1 are predicted to have clear histologic margins after resection. Patients with a PDS of 4 have unresectable tumors, and nonoperative palliation should be considered. Patients with a PDS of 2 or 3 have a high likelihood of positive histologic margins after resection and may be candidates for neoadjuvant chemotherapy.

A nonlethal conditioning approach to achieve durable multilineage mixed chimerism and tolerance across major, minor, and hematopoietic histocompatibility barriers.

Reconstitution of lethally irradiated mice with a mixture of syngeneic and allogeneic (A+B-->A) bone marrow results in multilineage mixed allogeneic chimerism, donor-specific transplantation tolerance, superior immunocompetence and resistance to graft-vs-host disease. However, the morbidity and mortality associated with lethal irradiation would be a major limitation to the clinical application of chimerism to induce tolerance for solid organ grafts or treat other nonmalignant hematologic diseases. We report here that durable multilineage mixed allogeneic chimerism and donor-specific transplantation tolerance for skin and primarily vascularized allografts can be achieved across multiple histocompatibility barriers using a nonmyeloablative radiation-based approach. The percentage of B10 mouse recipients that engrafted directly correlated with the degree of disparity between donor and recipient and the dose of total body irradiation administered. Although the occurrence of engraftment following conditioning with doses of total body irradiation of > or = 600 cGy was similar for animals receiving bone marrow disparate at MHC or MHC, minor and hematopoietic (Hh-1) loci (67% vs 78%), the level of donor chimerism was significantly less when multiple histocompatibility barriers were present (94.6 +/- 3.8% vs 37.5 +/- 12.5%). Treatment of the recipient with cyclophosphamide 2 days following allogeneic bone marrow transplantation reduced the dose of radiation sufficient for reliable engraftment to only 500 cGy of total body irradiation, regardless of MHC and Hh-1 disparity. Donor chimerism was stable and present in all lineages, with production of lymphoid (T and B cell), NK, and myeloid (erythrocyte, platelet, granulocyte, and macrophage) cells. Mixed chimeras exhibited donor-specific tolerance in vitro, as assessed by mixed lymphocyte culture (MLR) and cytotoxicity (CML) assays, and in vivo to skin and primarily vascularized cardiac allografts. The observation that engraftment and tolerance can be achieved across multiple histocompatibility barriers using nonmyeloablative recipient conditioning may allow allogeneic bone marrow transplantation to be applied to nonmalignant disease states in which lethal conditioning cannot be justified, including the induction of donor-specific tolerance for solid organ transplantation and the treatment of hemoglobinopathies and enzyme deficiency states.

Individual assessment of visual perception by surgeons observing the same laparoscopic organs with various imaging systems.

Laparoscopic surgery necessitates that operations be performed via a television screen; therefore, image quality is crucial in determining the final outcome. Electronic imaging systems are constantly undergoing revisions and incorporating new ideas. Recently, we have been confronted with a variety of systems, and we, as operators, have no idea what the differences are between the old and the newer systems. As clinicians, we have no electronic yardstick available to compare the options presented or to check the specifications. This study examines critical aspects of image perception. General surgeons, gynecologists, operating-room nurses, as well as allied health personnel all experienced in laparoscopic surgery participated as test subjects. A blinded study was performed in which various camera systems (single chip, three-chip, digitized and nondigitized) were displayed in random fashion. Porcine abdominal areas were displayed using the various imaging units. Participants graded image perception based on the following characteristics: sharpness, color, contrast, and depth of field. The audience were blinded to the types, brand, and number of cameras utilized. A significant preference was shown for digitized systems (P < 0.0001). Also, digitized single-chip cameras scored higher than three-chip cameras (P < 0.05). We propose thorough testing by surgeons and nursing personnel before deciding what type of equipment to purchase.

Phenotypic characterization of a novel bone marrow-derived cell that facilitates engraftment of allogeneic bone marrow stem cells.

Bone marrow transplantation is an accepted therapy for hematologic malignancies, aplastic anemia, metabolic disorders, and solid tumors. However, graft-versus-host disease (GVHD) and failure of engraftment have limited the widespread application of this technology to nonmalignant disease states. The use of purified bone marrow stem cells has been suggested as an approach to promote engraftment yet avoid GVHD. Although bone marrow stem cells, purified by cell sorting, engraft and repopulate lethally irradiated genetically identical recipients, they do not engraft in major histocompatibility complex (MHC)-disparate allogeneic recipients. We report for the first time the characterization of a novel cell population of donor bone marrow origin, separate from the hematopoietic stem cell, that facilitates engraftment of purified allogeneic bone marrow stem cells in an MHC-specific fashion without causing GVHD. Although 1,000 purified stem cells (c-kit+/Sca-1+/lineage-) reliably repopulate syngeneic mouse recipients, 10 times that number do not engraft in MHC-disparate allogeneic recipients. The addition of as few as 30,000 facilitating cells (CD8+/CD45R+/TCR-) is sufficient to permit engraftment of purified stem cells in MHC-disparate recipients. The cell surface phenotype of this purified cellular population differs significantly from other characterized lineages of lymphoid or myeloid origin. Based on multiparameter rare-events cell sorting, the facilitating fraction is CD8+, CD3+, CD45R+, Thy 1+, class IIdim/intermediate but alpha beta-TCR- and gamma delta-TCR-. This cellular population comprises approximately 0.4% of the total bone marrow and is separate from the hematopoietic stem cell. The coadministration of purified facilitating cells plus stem cells to optimize engraftment yet avoid GVHD may expand the potential application of bone marrow transplantation to disease states in which the morbidity and mortality associated with conventional BMT cannot be justified.

Evidence for early Th 2 T cell predominance in xenoreactivity.

Two distinct subsets of CD4+ Th lymphocytes have been characterized by their cytokine profiles: Th 1 (TH1) and Th 2 (TH2). While TH1 cells predominate in cell-mediated responses, TH2 cells support the humoral response. We have examined the mRNA cytokine profile of normal mouse lymphocytes in response to alloantigen versus xenoantigen (rat) in MLC, and present evidence to suggest that early in proliferative responses, alloreactivity is dominated primarily by TH1-type lymphocytes, while xenoreactivity is predominantly TH2. Normal mouse lymphocyte-responding cells were cultured in a one-way MLR with either allo or xeno antigen and examined for production of mRNA for cytokines characteristically produced by TH1 (IL-2, IFN-gamma) or TH2 (IL-4, IL-10) cells. Semiquantitative reverse transcription-polymerase chain reaction analysis was performed for mouse IL-2, IL-4, IL-10, and IFN-gamma mRNA. In the mouse anti-rat xeno response, mRNA for TH2 gene products were upregulated, with greater levels of IL-4 and IL-10 at 24 and 48 hr when compared with controls. In contrast, upregulation of mRNA for TH1 gene products occurred in the mouse anti-mouse allo response, with higher levels of IL-2 and IFN-gamma at 24 and 48 hr. In the anti-xeno response, upregulation of all 4 cytokines occurred by day 4 and peak levels of mRNA for all cytokines examined were 2-3 times that seen for the peak anti-allogeneic response. These data suggest that early xenorecognition may differ from allorecognition by differential activation of the TH2 subset. A better understanding of the balance between Th subset function and cytokine profile in allo and xeno reactivity may allow a more targeted and specific approach to control the early events in xenograft rejection.

Characterization of the immunosuppressive effects of nitric oxide in graft vs host disease.

The generation of nitric oxide (.N = O) during in vitro assays involving lymphocyte-macrophage interaction can result in profound inhibition of lymphocyte proliferation. The present study examined whether .N = O synthesis plays a role in the suppression observed in immune function assays during graft vs host disease (GvHD). By using a parent to F1 model to induce GvHD (C57BL/6J to C57BL/6J x DBA 2J F1), a mild but transient increase in serum NO2- plus NO3- levels was observed on day 12 after inoculation. Resident peritoneal macrophages obtained from mice with GvHD demonstrated enhanced .N = O synthesis in response to LPS, compared with control F1 peritoneal macrophages. Similarly, when splenocytes from GvHD mice were cultured with Con A or LPS enhanced supernatant NO2- levels were observed, compared with control F1 mice. Addition of NG-monomethyl-L-arginine (NMA), a competitive inhibitor of .N = O synthesis, resulted in decreased NO2- levels and greatly enhanced proliferation in response to Con A. Addition of NMA to LPS-stimulated cultures did not enhance proliferation, perhaps as the result of the paucity of B cells in the GvHD population. LPS-induced .N = O synthesis by GvHD splenocytes was blocked by anti-IFN-gamma mAb, whereas Con A-induced .N = O synthesis was relatively unaffected by similar concentrations of anti-IFN-gamma mAb, suggesting different mechanisms of induction of .N = O synthesis. A proliferative response of splenocytes from mice with GvHD to third-party alloantigen was not detectable, even in the presence of NMA. The suppression observed when splenocytes from GvHD animals were added to control TNP-modified self cultures was partially reversed in the presence of NMA. These results demonstrate that .N = O synthesis in both splenocyte and peritoneal macrophage populations from GvHD mice is enhanced, revealing that in vivo priming of macrophages for .N = O synthesis occurs during GvHD. Some, but not all, in vitro tests of immune function by using GvHD splenocytes are suppressed by the generation of .N = O.

CD4+ T cells, but not CD8+ T cells, mediate the breaking of tolerance in mixed allogeneic chimeras (B10 + B10.BR-->B10).

Reconstitution of mouse recipients with a mixture of syngeneic plus allogeneic bone marrow (A+B-->A) results in stable mixed lymphohematopoietic chimerism and donor-specific transplantation tolerance. Previously, it was reported that administration of large numbers of unmanipulated host-type splenocytes to neonatal or adult radiation bone marrow chimeras resulted in a loss of chimerism and donor-specific transplantation tolerance. To characterize the phenotype(s) of cells that were responsible for this loss of chimerism, we performed depletion of various subsets of unmanipulated B10 splenocytes prior to infusion into mixed allogeneic chimeras (B10 + B10.BR-->B10). Recipients were followed serially to identify changes in the level of donor chimerism and by in vitro functional assays of tolerance. We report here that CD4+ T cells, but not CD8+ T cells, were sufficient to mediate the loss of donor chimerism. In all recipients in which allogeneic chimerism became undetectable, there was a simultaneous loss of donor-specific transplantation tolerance.