Cyclosporin A-sensitive decrease in the transmembrane potential across the inner membrane of liver mitochondria induced by low concentrations of fatty acids and Ca2+.

At low Ca2+ concentrations the pore of the inner mitochondrial membrane can open in substates with lower permeability (Hunter, D. R., and Haworth, R. A. (1979) Arch. Biochem. Biophys., 195, 468-477). Recently, we showed that Ca2+ loading of mitochondria augments the cyclosporin A-dependent decrease in transmembrane potential (DeltaPsi) across the inner mitochondrial membrane caused by 10 micro M myristic acid but does not affect the stimulation of respiration by this fatty acid. We have proposed that in our experiments the pore opened in a substate with lower permeability rather than in the "classic" state (Bodrova, M. E., et al. (2000) IUBMB Life, 50, 189-194). Here we show that under conditions lowering the probability of "classic pore" opening in Ca2+-loaded mitochondria myristic acid induces the cyclosporin A-sensitive DeltaPsi decrease and mitochondrial swelling more effectively than uncoupler SF6847 does, though their protonophoric activities are equal. In the absence of P(i) and presence of succinate and rotenone (with or without glutamate) cyclosporin A either reversed or only stopped DeltaPsi decrease induced by 5 micro M myristic acid and 5 micro M Ca2+. In the last case nigericin, when added after cyclosporin A, reversed the DeltaPsi decrease, and the following addition of EGTA produced only a weak (if any) DeltaPsi increase. In P(i)-containing medium (in the presence of glutamate and malate) cyclosporin A reversed the DeltaPsi decrease. These data show that the cyclosporin A-sensitive decrease in DeltaPsi by low concentrations of fatty acids and Ca2+ cannot be explained by specific uncoupling effect of fatty acid. We propose that: 1) low concentrations of Ca2+ and fatty acid induce the pore opening in a substate with a selective cation permeability, and the cyclosporin A-sensitive DeltaPsi decrease results from a conversion of DeltaPsi to pH gradient due to the electrogenic cation transport in mitochondria; 2) the ADP/ATP-antiporter is involved in this process; 3) higher efficiency of fatty acid compared to SF6847 in the Ca2+-dependent pore opening seems to be due to its interaction with the nucleotide-binding site of the ADP/ATP-antiporter and higher affinity of fatty acids to cations.

Acute abdomen with colonic necrosis induced by Kayexalate-sorbitol.

Colonic necrosis is an unusual complication after treatment of hyperkalemia with sodium polystyrene sulfonate (SPS, Kayexalate) in sorbitol. To increase awareness of this complication, we report a case of necrosis of the transverse colon in a patient given oral and rectal SPS-sorbitol for hyperkalemia. Colonic necrosis was manifested as an acute abdomen within 24 hours of initial administration. Prompt surgical resection of the necrotic transverse colon permitted rapid recovery of bowel function. Although SPS crystals are seen microscopically in the necrotic bowel, experimental evidence implicates the sorbitol component of the SPS-sorbitol in the pathogenesis of colonic necrosis. A high index of suspicion for the unusual complication of colonic necrosis after oral or rectal administration of SPS-sorbitol may allow prompt recognition and surgical cure.

Sepsis impairs gut amino acid absorption.

Sepsis has been shown to adversely affect the barrier and metabolic functions of the small intestine as well as to reduce mesenteric blood flow and cause histologic damage. However, the effect of sepsis on gut absorptive function has been largely ignored. In this study, intestinal absorption of arginine and an amino acid analogue, aminoisobutyric acid, was studied using in vivo and in vitro techniques in an experimental model of sepsis. In vivo studies showed a significant impairment in the absorption of both amino acids from the intestinal lumen 24 and 72 hours after cecal ligation and puncture. Uptake of these amino acids by everted gut sacs prepared from septic animals was also significantly reduced. This reduction in absorptive capacity of the gut may limit the ability of enteral feeding alone to supply nutritional requirements during sepsis and may also contribute to the associated morbidity and mortality.

Arginine enhances wound healing and lymphocyte immune responses in humans.

Arginine has been shown to enhance wound healing and T-cell-mediated immune function in rodents. In this study the effect of oral arginine supplementation on human collagen synthesis and T-cell function was studied in 36 healthy, nonsmoking human volunteers. While volunteers were under local anesthesia, a 5 cm segment of expanded polytetrafluoroethylene tubing (1 mm outer diameter, 90 mu pore size) was inserted subcutaneously into the right deltoid region. The volunteers were then randomized into three groups that were given the following substances: (1) daily supplements of 30 gm arginine hydrochloride (24.8 gm free arginine); (2) 30 gm arginine aspartate (17 gm free arginine) daily; or (3) placebo. The supplements were given orally for 2 weeks; dietary intake was not controlled. Mitogenic responses of peripheral blood lymphocytes to phytohemagglutinin and concanavalin A were assayed at the start of study and at 1 and 2 weeks after supplementation. At 2 weeks the catheters were removed, and the amount of hydroxyproline was determined as an index of new collagen synthesis and deposition. Arginine supplementation significantly enhanced the amount of collagen deposited into a standardized wound as assessed by the amount of hydroxyproline present (10.1 +/- 2.32 nmol/cm graft in controls vs 17.57 +/- 2.16 nmol/cm in the arginine aspartate group, [p = 0.028] and vs 23.85 +/- 2.16 nmol/cm in the arginine hydrochloride group [p less than 0.001]). In parallel, arginine supplementation at both doses increased lymphocyte mitogenesis in response to phytohemagglutinin and concanavalin A. The data suggest that arginine may be of clinical benefit in improving wound healing and immune responses.

The use of tissue expanders in immediate breast reconstruction following mastectomy for cancer.

From April 1985 to March 1987, 50 women had 56 mastectomies for documented cancer. A tissue expander was used for immediate reconstruction in each case. The patient records were used to determine morbidity and mortality, as well as to examine the effect of reconstruction on adjuvant therapy, cancer surveillance, adequacy of cancer surgery and patient satisfaction. The patients were followed up for an average of 13 months. Fifty-two of 56 expanders were successfully replaced with a permanent prosthesis. Forty-seven women remain alive. Local or regional recurrence did not occur in any women within the study period. Complications, of which superficial skin necrosis was most common, occurred in 35% of reconstructions and were investigated critically. Patient satisfaction was high. Dissatisfaction was not related to the incidence of complications but rather appeared to reflect the success of the patient's cancer treatment.

The wound is a possible source of posttraumatic immunosuppression.

Wound fluid from 10-day-old healing wounds in rats inhibits lymphocyte immune responses. Since severe injury is frequently complicated by immunosuppression as manifested by sepsis, we hypothesized that the wound may be a source of factors that impair host immune responses. Therefore, we studied the effect of systemic wound fluid administration on the survival of rats subjected to an acute peritonitis model. Male Sprague-Dawley rats, fitted with internal jugular catheters 48 hours previously, underwent cecal ligation and puncture with a 23-gauge needle. Immediately after the operation, rats were treated intravenously every 12 hours with either wound fluid obtained from 10-day-old healing wounds and adjusted to 10 mg of protein per milliliter or rat serum. In vitro testing of the wound fluid showed it to be highly inhibitory of thymic lymphocyte mitogenesis. Rats treated with wound fluid had significantly higher mortality after peritonitis than did control rats. The data show that the wound contains factors that can impair host immune responses to sepsis. This suggests that the wound may be the source of posttraumatic host immunosuppression.

Trends and controversies in the management of carcinoma of the stomach.

The marked decrease in the incidence of carcinoma of the stomach during the past four decades has been worldwide, with a few exceptions. Dietary deficiencies or carcinogenic substances appear to be the most important environmental factor in the genesis of carcinoma of the stomach. The risk of carcinoma of the stomach appears to be determined early in life. Based on the histologic features and the distinct differences in clinical behavior, adenocarcinoma of the stomach can be divided into the intestinal and diffuse types of lesions. In high risk populations, such as the Japanese, the intestinal type is found more frequently, while white Americans tend to have diffuse carcinomas. Premalignant gastric lesions, such as adenomatous polyps, are uncommon causes of carcinoma. The symptoms of carcinoma of the stomach are not specific and there is no satisfactory noninvasive diagnostic test to detect the carcinoma early. In Japan, mass screening, using air contrast radiography and endoscopy of the upper part of the gastrointestinal tract with biopsies and cytologic sampling, leads to detection of early stages of carcinoma of the stomach in 40 per cent of patients. The postsurgical survival rate of these patients is significantly higher, compared with those who have advanced disease. Mass screening for carcinoma of the stomach would not be cost effective in North America. The only curative treatment for carcinoma of the stomach is resection, which is most successful in early stages of carcinomas of the stomach. Radical subtotal gastrectomy is the operation of choice for curative treatment of carcinomas of the body and antrum of the stomach. In Japan, a more extensive lymphadenectomy has improved the five year survival rate. Operations for carcinoma of the cardia are invariably palliative. Palliative resection of an incurable carcinoma of the stomach has been shown to be more worthwhile than a bypass procedure. Adjuvant chemotherapy seems to have improved the unsatisfactory results of radical resection in the treatment of locally advanced carcinoma in Japan, a finding that has not been confirmed in the United States or Europe. The use of intraoperative electron beam radiation to improve the surgical outcome of carcinoma of the stomach is being investigated.

The effect of in vivo T helper and T suppressor lymphocyte depletion on wound healing.

The role of T lymphocytes in wound healing is still not well-defined. Because it had been previously shown that in vivo depletion of T cells leads to impaired wound healing, the effect of depleting T cell subsets on subsequent fibroplasia was studied. T helper/effector cells were depleted by the use of the monoclonal antibody GK1.5, reactive against the L3T4 antigen (CD4). T suppressor/cytotoxic lymphocytes were depleted by using the 2.43 monoclonal antibody reactive against the Lyt 2 antigen (CD8). In the first experiment, Balb/c mice were treated with the antibodies starting at 24 hours before wounding was performed, and weekly thereafter. Depletion of the T helper/effector cells had no effect on wound-breaking strength or hydroxyproline deposition in sponge granulomas, whereas depletion of T suppressor/cytotoxic cells significantly enhanced both of these healing parameters. In a second experiment, T cell subset depletion was started on Days 0, 3, 7, 10, and 14 postwounding, and treatments were continued weekly thereafter. Once again, depletion of T helper/effector cells had no effect on wound healing, whereas depletion of T suppressor/cytotoxic cells markedly increased both wound-breaking strength and collagen synthesis. In conclusion, the data show that T suppressor/cytotoxic cells have a counter-regulatory role in wound healing, whereas the T cell subset responsible for up-regulating wound healing remains to be identified.

Stimulation of T cell immunity by arginine enhances survival in peritonitis.

T cell-mediated immunity may play a role in host responses to infection. Arginine is a known thymic and T cell stimulator which enhances host allogenic, mitogenic, and anti-tumor responses. We, therefore, examined the effect of arginine on the survival of rats with severe and lethal peritonitis induced by cecal ligation and double-needle puncture (CLP). In Experiment 1, arginine HCl (100 mg) was given bid by gavage starting immediately after CLP. In Experiment 2, the same dose of arginine was given by gavage bid for 3 days pre-CLP and continued thereafter. In Experiment 3, arginine was administered iv post-CLP (100 mg tid). Arginine had no effect on overall survival in Experiment 1. In Experiments 2 and 3, arginine therapy significantly increased survival at all times. A separate experiment was carried out to determine the reason for the differential response to arginine administered via gavage or iv post-CLP (Experiments 1 and 3). Nonseptic rats showed a 400% increase in plasma arginine 30 min after gavage with 100 mg arginine (P less than 0.001). No rise in plasma arginine was noted when arginine was administered by gavage post-CLP. The impaired intestinal absorption or markedly increased utilization of arginine in this septic model may explain why no improved survival was seen in Experiment 1. The mechanism for the improved survival with arginine therapy seen in Experiments 2 and 3 may be related to its known thymic and T cell immunostimulatory effects.

Suppressor cell generation during normal wound healing.

We have previously shown that 10 day healing wounds in rats contain wound mononuclear cells (WMNC) which inhibit normal lymphocyte mitogenic and allogeneic responses. In the present study we sought to further characterize the WMNC and define their mechanism of action. Polyvinyl alcohol sponges implanted in wounds were harvested and processed 10 days postwounding. The resultant WMNC suspension contained less than 15% macrophages. By FACS analysis, 69.5 +/- 11.4% (mean +/- SD of eight separate experiments) of the cells expressed the all T cell marker (W3/13), while 47.7 +/- 11.9% stained with the T helper/effector marker (W3/25) and 49.5 +/- 18.8% expressed the T suppressor/cytotoxic phenotype (OX8) (Th/Ts ratio = 0.96 +/- 0.13). When various numbers of WMNC were cocultured with 5 X 10(5) PHA-stimulated rat thymic lymphocytes, as few as 500 WMNC inhibit normal blastogenesis. Long-term (72 or 144 hr) culture of WMNC revealed that they maintain their suppressive activity. Furthermore, the conditioned media of long-term cultures also significantly suppressed thymic lymphocyte PHA blastogenesis, suggesting that the WMNC secrete suppressive cytokines. Large doses of human recombinant IL-2 or indomethacin did not abrogate the inhibitory effect of WMNC. We conclude that the healing wound is normally infiltrated by suppressor lymphocytes which generate immune inhibitory cytokines.

Generation of an anti-interleukin 2 factor in healing wounds.

Previously we have noted that fluid obtained from ten-day-old healing wounds noncytotoxically inhibits the blastogenesis of lymphocytes in response to mitogens or antigens. Since these lymphocytic responses are interleukin 2 (IL-2)-mediated, we looked for a specific IL-2 inhibitor in wound fluid. We have found that wound fluid blocks the response of thymic lymphocytes and of two cloned T-helper cell lines (D10 and HT2) to exogenous human recombinant IL-2. The wound fluid enhances fibroblast proliferation, thus demonstrating that its proliferative inhibitory activity is specific for lymphocytes. The findings suggest that wound fluid contains a factor that impairs lymphocyte response to IL-2, probably at the receptor or postreceptor level.

Significance of T-lymphocytes in wound healing.

To determine the importance of T-lymphocytes in wound healing, we examined the effect of T-lymphocyte depletion on the healing of surgical wounds. Thirty Balb/c mice were injected intraperitoneally with 1 mg of rat anti-mouse (IgG2b) cytotoxic monoclonal antibody (30H12) against the Thy1.2 (all T) determinant. Twenty-four hours later animals showed a greater than 95% depletion of Thy1.2 cells in peripheral blood and spleen. Thirty control mice received nonspecific rat immunoglobulin (1 mg). Twenty-four hours after treatment mice underwent a 2.5 cm dorsal skin incision with subcutaneous placement of polyvinyl alcohol sponges. Injections were repeated at weekly intervals. Wound healing was assessed at 2, 3, and 4 weeks by the breaking strength of wound strips and by the hydroxyproline content of sponge granulomas (an index of wound reparative collagen deposition). Thy1.2 depletion at death was 95% to 57% in peripheral blood and 86% to 68% in the spleen. Both groups gained weight equally. We found that T cell depletion significantly impairs wound breaking strength and wound collagen deposition at all times studied. The data strongly suggest that T-lymphocytes modulate fibroblast activity during normal wound healing.

Lymphocyte participation in wound healing. Morphologic assessment using monoclonal antibodies.

To investigate lymphocyte participation in wound healing, the migration of T lymphocyte subsets into healing wounds and subcutaneously implanted polyvinyl alcohol sponges was studied. Frozen sections of 5-, 7-, and 10-day-old incisional wounds and sponges from Lewis rats were stained with mouse anti-rat monoclonal antibodies. Cellular staining to OX1 (all leucocyte), W3/25 (helper/effector T lymphocytes), and OX8 (suppressor/cytotoxic T lymphocytes) was quantitated in two arbitrarily defined areas based on maximal cellular infiltration: the superficial wound, down to and including the papillary dermis, and the deep wound, the reticular dermis. Five-day wounds were significantly more cellular than 10-day wounds in the deep portion (p less than 0.05) and somewhat more cellular in the superficial section (p less than 0.10). Approximately 2:1 W3/25 to OX8 ratios were noted for wound strips on all days. At 5 and 10 days there are twice as many W3/25 and OX8 labeled cells in the deep wound as in the superficial portion. At 7 days there is a peak in surface W3/25 and OX8 lymphocytes, whereas the deep population remains constant. Seven- and 10-day sponge granulomas demonstrate ratios similar to the wound strips (5-day sponge lymphocytic infiltration was insufficient to count). The data demonstrate that lymphocyte subpopulation participation in wound healing is a dynamic and distinctive process.

Current concepts and controversies concerning the etiology, pathogenesis, diagnosis, and treatment of malignant tumors of the anus.

Malignant disease of the anus is uncommon. Possible predisposing causes include chronic inflammation and a transmissible agent. Epidemiologic studies suggest an increased incidence in homosexuals. With the exception of mucoepidermoid and small-cell carcinoma, the morphology of anal carcinoma has little influence on treatment and prognosis. Site, size of the primary lesion, and the presence of groin metastases are the crucial factors in prognosis. There is no satisfactory method for staging anal carcinoma--the symptoms are nonspecific. Diagnosis is based on histologic examination of biopsy material or tissue obtained from anal operations. The treatment of infiltrating, recurrent, or residual malignant anal lesions is a radical abdominoperineal resection. The addition of a limited obturator and hypogastric lymphadenectomy may be worthwhile. Inguinal lymphadenectomy provides palliation in the treatment of synchronous groin metastases, whereas in cases of metachronous metastases, groin dissection may result in an occasional cure. Small, noninfiltrating, low-grade anal lesions are best treated by either adequate local excision or supervoltage radiotherapy. If borne out, the promising results obtained with the combined chemoradiotherapy for anal cancer followed by local excision of the residuum will radically alter the future management of carcinoma of the anus.

Interleukin 2 enhances wound healing in rats.

Antigen-stimulated lymphocytes secrete lymphokines which have been shown to enhance in vitro fibroblast migration, proliferation, and protein synthesis. In the present experiments, the effect of human recombinant interleukin 2 (RIL-2) on wound healing was assessed in vivo. Groups of male Lewis rats, 225-250 g, underwent intraperitoneal insertion of osmotic pumps and a 7-cm dorsal skin incision with subcutaneous placement of polyvinyl alcohol sponges under anesthesia. The dorsal wounds were closed with stainless-steel sutures. The dose of RIL-2 administered was 60,000 u/rat/day for 7 days in experiment I, and 140,000 u/rat/day for 7 days in experiment II. Controls received equal volumes of excipient. Animals were sacrificed 10 days post wounding and wound healing was assessed by fresh breaking strength, fixed breaking strength (following 72 hr of Formalin fixation which maximally crosslinks the collagen present), and sponge hydroxyproline content (an index of reparative collagen accumulation). In vivo RIL-2 administration significantly augmented wound fresh and fixed breaking strength and wound collagen synthesis. Higher doses of RIL-2 (experiment II) did not result in further increases in the parameters studied. The data suggest that lymphocytes participate directly in the process of wound healing.

The diagnostic value of computed tomography in a selected case of penetrating thoraco-abdominal injury.

A patient with a penetrating thoraco-abdominal injury produced by a pencil left in situ is described. CT scan accurately demonstrated the innocuous path of the pencil which was removed without sequelae. CT scan may be valuable in the management of penetrating injuries of the trunk with weapon still in place.

Transhiatal (blunt) esophagectomy for malignant and benign esophageal disease: clinical experience and technique.

"Blunt" transhiatal esophagectomy was performed in 23 selected patients. Nineteen had squamous carcinoma of the esophagus (upper third, 1; middle third, 12; distal third, 6), and 2 had adenocarcinoma of the distal esophagus. The other 2 patients had severe lye strictures. Resection with reconstruction was performed in one stage. Esophagogastric continuity was restored using the stomach in the posterior mediastinal position in 20 patients and in the substernal position in 2. The colon in the posterior mediastinal position was used in 1 patient with a lye stricture. Transmural tumor extension or cervical or celiac nodal metastases or both were present in 18 of 21 patients with carcinoma. There was 1 hospital death due to pericardial tamponade. Morbidity included a transient cervical anastomotic leak in 3 patients, one temporary and three permanent unilateral recurrent laryngeal nerve palsies, one intraoperative splenic injury, and severe hemorrhage requiring sternotomy for control in 1 patient. Pulmonary complications occurred in 4 patients: aspiration pneumonia (1) and moderate atelectasis (3). Three patients have died (11, 12, and 17 months postoperatively) in the group with cancer, with follow-up time of 3 to 30 months (mean, 15 months). Transhiatal blunt esophagectomy is a safe and effective procedure in many patients with either esophageal cancer or extensive, benign esophageal strictures.

Intravenous hyperalimentation with high arginine levels improves wound healing and immune function.

The purpose of this study was to evaluate the effect of increased arginine levels in intravenous hyperalimentation (IVH) therapy on wound healing and thymic immune function. Groups of SD rats, 275-325 g, underwent placement of internal jugular catheter, 7-cm dorsal skin wounding, insertion of polyvinyl alcohol sponges subcutaneously, and closure of wounds with stainless-steel sutures. Twenty-four hours later, rats were started on IVH at a rate of 0.8-1 ml/100 g body wt/hr. All IVH solutions contained 20% dextrose, adequate amounts of minerals and vitamins, and two different amino acid mixtures: (A) Fre III (4.05 g ARG/liter) (n = 13); (B) experimental (7.50 g ARG/liter) (n = 11). Solutions were isonitrogenous, and contained similar amounts of essential amino acids. After 7 days of IVH, weight gain did not differ between the two groups; however, cumulative N balance was superior in group A. Wound healing was improved in group B as assessed by fresh wound strip breaking strength, fixed breaking strength, and the amount of reparative collagen deposition as assessed by the hydroxyproline content of the implanted sponges. Group B animals also had improved thymic function as assessed by thymic weight, the total number of thymic lymphocytes/gland and mitogenic reactivity of thymic lymphocytes to PHA and Con A. The experiments indicate that high arginine levels in IVH solutions improve wound healing and thymic immune function following injury.

Cocaine colitis. Is this a new syndrome?

An unusual case of colitis in a 37-year-old cocaine addict is described. The patient presented with right-sided abdominal pain and diarrhea exacerbated by his use of cocaine. Significant antibiotic ingestion was denied. At laparotomy, an edematous cecum and ascending colon were found, the cut surface of which revealed diffuse superficial ulcerations and yellowish fibrinous material. Microscopic examination demonstrated findings consistent with pseudomembranous colitis with an ischemic component. A mechanism involving catecholamine-induced mucosal ischemia is postulated to explain the findings seen in this patient.

Inhibition of host immunity by fluid and mononuclear cells from healing wounds.

Severe trauma impairs host immunity, which in turn renders the host susceptible to infection often terminating in death. This impairment occurs 7 to 14 days after injury, a time when wound healing is at its maximum. We examined the interactions of wound healing to host immunity by studying the in vitro and in vivo immune effects of wound components (i.e., wound fluid [WF] and wound mononuclear cells [WMNC]). Lewis male rats (RT-1(1] weighing 300 to 350 gm underwent 7 cm dorsal skin incisions and subcutaneous placement of polyvinyl alcohol sponges. At 7 and 10 days after wounding, sponges were removed and WF was separated from the cellular elements. The cell suspension was purified to contain 80% to 90% WMNC. Ten percent WF from 7- and 10-day-old wounds inhibits normal thymic lymphocyte blastogenesis to concanavalin A and phytohemagglutinin. Addition of 5 X 10(4) WMNC leads to similar inhibition. WF and WMNC from 10-day-old wounds also inhibit in vitro allogeneic responses tested in one way MLR of Lewis splenocytes with inactivated ACI (RT-1a) spleen cells by 75% to 96% and 85% to 98%, respectively. The inhibitory action of WF is heat resistant (56 degrees C for 30 minutes) and noncytotoxic. In vivo allogeneic responses, tested by grafting ACI skin onto Lewis recipients, were inhibited by intraperitoneal administration of 10-day-old WF (p less than 0.01). We conclude that WF contains factor(s) that inhibit in vitro and in vivo immune responses. WMNC exhibits the same action, suggesting that they may be the source of the WF inhibitory factor(s). These findings may explain host immunosuppression after severe trauma.