Survival following orthotopic cardiac xenotransplantation between juvenile baboon recipients and concordant and discordant donor species: foundation for clinical trials

It has been more than a decade since the last clinical trial of cardiac xenotransplantation in a newborn infant. Since that event, laboratory research at Loma Linda University has focused on survival studies of orthotopically xenografted juvenile baboon recipients. Both concordant and discordant donor species have been used. Transgenic donors have not been explored at Loma Linda. Instead, simplified host immunoregulative protocols, consistent with those used in neonatal cardiac allografting, have been adapted to xenotransplant research. Xenograft bridge to alloengraftment was evaluated in a series of five juvenile baboon recipients. Heterotopically implanted cardiac xenografts stimulated host production of xenoreactive antibody. Orthotopic cardiac allografting was then carried out. Xenoantibody appeared to play little role in immediate or chronic survival of experimental hosts. A clinical protocol of xenobridging to allotransplantation would likely succeed. Two consecutive series of orthotopically xenotransplanted hosts using rhesus monkey cardiac donors demonstrated unprecedented long-term survival. Splenectomy combined with maintenance therapy consisting of FK-506 and methotrexate contributed to survival of up to 502 days in one series of xenografted baboon hosts selected for ABO blood grouping, mixed lymphocyte culture, and crossmatch compatibility. Survival beyond a year (maximum 515 days) among three consecutive juvenile baboon recipients of orthotopically implanted rhesus monkey hearts, in which splenectomy was omitted and cyclosporine was substituted for FK-506, represents a benchmark achievement. Commencing maintenance immunosuppression several weeks prior to transplantation appeared to improve chronic survival significantly. Investigation of discordant (pig-to-baboon) host survival has focused on adsorption of naturally occurring xenoreactive antibody at the time of transplantation. This strategy, combined with pretransplant total lymphoid irradiation and both pre- and posttransplant immunosuppression, succeeded in preventing hyperacute rejection and resulted in survival of up to 24 days, thereby permitting observation of the delayed xenograft rejection phase. Data support consideration of additional clinical trials of concordant neonatal cardiac xenotransplantation and offer promise for the development of discordant xenotransplantation as an ultimate therapeutic resource.     

Vesnarinone prolongs survival and reduces lethality in a murine model of lethal endotoxemia

Vesnarinone (3,4-Dihydro-6-[4(3,4-dimethoxybenzoyl)-1-piperanizyl]-2(1H)-quino linone), a recently synthesized quinolinone derivative with positive inotropic properties, has been reported the survival of patients with chronic congestive heart failure. However, the mechanisms that contribute to this improvement are not yet well understood. There is increasing evidence that vesnarinone has novel immunosuppressive properties related to its inhibition of cytokine production. Cytokines have been shown to play a pivotal role in the pathophysiologic consequences of fatal bacteremic shock. In this study, we investigated the effects of vesnarinone in a murine model of lethal endotoxemia induced by lipopolysaccharide (LPS). Eight-week-old female BALB/c mice were given 300 or 400 micrograms of LPS, and 50 or 100 mg/kg of vesnarinone was administered by oral gavage and/or 10 or 30 micrograms of vesnarinone was given intra peritoneally. Vesnarinone prolonged the median survival time and reduced lethality when given at the same time as the LPS injection. However, vesnarinone did not have a beneficial effect when administered 2 hours after LPS treatment. Plasma TNF-alpha reached a maximum level 1 hour after LPS challenge, and vesnarinone reduced the plasma level of TNF-alpha, when administered at the same time as LPS injection. Vesnarinone had protective effects against lethal endotoxemia; these effects were considered to be due to the suppression of TNF-alpha production. These findings suggest that vesnarinone may be a promising agent for the treatment of bacterial sepsis and shock.     

Triple immunosuppression protects murine intracerebral, hippocampal xenografts in adult rat hosts: effects on cellular infiltration, major histocompatibility complex antigen induction and blood-brain barrier leakage

Recently we reported protection of intracerebral mouse to rat hippocampal xenografts upon treatment with a combination of cyclosporin A, prednisolone and azathioprine. These findings are now supported in an extended analysis of graft-infiltrating cells. Host T-cell and macrophage infiltration and the immunocytochemical level of cellular expression of major histocompatibility complex class I and II antigens, measured by densitometric analysis, were compared between recipient rats receiving cyclosporin A alone or cyclosporin A in combination with prednisolone and azathioprine. The combination therapy resulted in a much improved survival of the xenografted hippocampal tissue with preservation of organotypic granule and pyramidal cell layers. Graft infiltration by T-cells and macrophages was significantly lower and the level of major histocompatibility complex class I and II antigen expression by the infiltrating cells markedly reduced. Lower expression of donor-type major histocompatibility complex class I antigen was also found in the xenografts in the trimedicated recipients, together with reduced blood brain barrier leakage and astrogliosis at the host-graft interface. The results demonstrate the benefits of using combined immunosuppressive strategies for protection of histoincompatible brain xenografts in the central nervous system.     

Highly sensitive and specific polymerase chain reaction assays for detection of baboon and pig cells following xenotransplantation in humans

Pig and baboon xenotransplants in humans require assays that discriminate source from human cells to investigate engraftment and identify true infection of recipients with xenogeneic endogenous retroviruses. We developed two polymerase chain reaction assays that target a porcine-specific sequence in the beta-globin gene and a baboon-specific sequence in the mitochondrial cytochrome oxidase subunit II gene. The sensitivity and specificity of both assays were evaluated on DNA lysates from baboon, pig, and human peripheral blood lymphocytes. Both assays detected single cells in backgrounds of human DNA. Additionally, both assays were highly specific and yielded negative results in reactions containing only human DNA. The two assays reliably detected peripheral blood lymphocyte samples from 14 baboons and 16 pigs. The baboon- and pig-specific target sequences are gender independent and nonpolymorphic and allow universal applicability. The high sensitivity and specificity is of particular importance in assessing low-level engraftment in human xenotransplant chimeras.     

T cell-mediated induction of airway hyperresponsiveness and altered lung functions in mice are independent of increased vascular permeability and mononuclear cell infiltration

To evaluate extra-natriuretic effects of atrial natriuretic peptide (ANP), plasma ANP (pANP) levels were assessed in seven healthy men on low-sodium diet (80 mEq NaCl/day), in basal conditions and during stepwise infusion of human ANP (2, 4, 8 and 16 ng/min/kg). To determine the individual physiological (PHY) pANP level, we measured pANP in the same subjects after a high-salt diet (400 mEq NaCl/day), that is, in a physiological stimulation of ANP. We then compared the effects of the PHY levels of ANP to the effects of pharmacological (PHA) pANP levels. Neither PHY nor PHA pANP levels modified creatinine clearance or blood pressure. The progressive rise in pANP levels was associated with increases in urinary excretion of Na+, K+ and urea. ANP alone respectively accounted for 41%, 30% and 92% of the increase in natriuresis, kaliuresis and urea excretion that occurred after changing salt intake from 80 to 400 mEq/day. Pharmacological ANP levels raised CH2O and reduced UOsm. Interestingly, PHA levels were associated with significant decrease in serum K+ (from 4.5 +/- .1 to 4.0 +/- .1 mEq/liter) and plasma urea (from 31.9 +/- 5 to 24.2 +/- 4 mg/dl). The mean cumulative urinary potassium and urea losses corresponded to the theoretical body losses of potassium and urea; moreover, the individual cumulative urinary losses of potassium and urea significantly correlated with the corresponding decrement in their plasma levels. In conclusion, ANP has both physiological and pharmacological significance in the control of potassium and urea metabolism by decreasing plasma levels of K+ and urea through effects on the renal excretory function.     

Monoclonal antibody to the HLA class I alpha 3 domain inhibits T cell activation and prolongs cardiac allograft survival in HLA-transgenic mice

Workload measurement is one of the most reliable tools in perspective payment systems, to evaluate clerk and technical personnel needs in a Surgical Pathology Department. In 1992 Italian Government began a full-comprehensive modification of financial support in National Health Care System, and during the last four years Public Hospitals and Local Health Units modified their financial organization with the introduction of workload measurement for the personnel needs. We propose a synthetic method for the workload measurement in Surgical Pathology Departments, similar to that proposed for perspective payment of surgical pathology services. The method is reliable and simple representing the full performance to obtain pathological report.     

Castanospermine, an oligosaccharide processing inhibitor, reduces membrane expression of adhesion molecules and prolongs heart allograft survival in rats

The clinical characteristics and natural history of 55 cases with antenatally diagnosed fetal uropathy were investigated. Percutaneous aspiration of the fetal pelvic or vesical urine was performed to decompress progressive unilateral hydronephrosis in 2 cases and to evaluate renal function in another 2 cases of bilateral hydronephrosis. As the postnatal diagnosis, upper urinary tract dilatation (hydronephrosis or hydronephroureter, 33 cases) and renal dysplasia (15 cases) made up 87% of all cases. A combination of hydronephrosis in one kidney and renal dysplasia in the other was also found in another 2 cases. Among 35 cases with upper urinary tract dilatation, 27 cases demonstrated pelviureteric junction stenosis and surgical intervention was necessary in 15 cases. In 17 cases with renal dysplasia, spontaneous regression was observed in only 3 cases and surgical intervention by means of percutaneous nephrostomy and nephrectomy was performed in 4 and 6 cases, respectively.     

Peripheral blood granulocytes and mononuclear cell responses in monkeys with experimental shigellosis

Between January 1992 and November 1992, four consecutive patients (ages 53 to 81 years) underwent early surgical repair of postinfarction ventricular septal ruptures using a new simple operative technique. The principles of the technique are longitudinal incision of the infarcted left anterior ventricular wall, placement of a saccular patch of single equine pericardium that covers the infarcted left ventricular wall, and large buttressed suture closure of the left ventriculotomy. The infarcted septum and infarcted left ventricular wall are completely separated from the left ventricular cavity. In this procedure, the infarcted myocardium is not resected, and left and right ventricular muscles are preserved. This technique is simple and safe for use in the acute phase of myocardial infarction, and it preserves ventricular function after surgery.     

Angiotensin II receptor antagonist TCV-116 reduces graft coronary artery disease and preserves graft status in a murine model. A comparative study with captopril

BACKGROUND: Despite the current progress in immunosuppressive regimens, the incidence of graft coronary artery disease (CAD) after cardiac transplantation has not decreased. Recent study has revealed that angiotensin-converting enzyme (ACE) inhibition decreases CAD in rats; however, it is not clear whether this beneficial effect of ACE inhibition is due to a decrease in production of angiotensin II (Ang II) or inhibition of bradykinin degradation. To determine whether Ang II type 1 receptor (AT1-R) blockade has an inhibitory effect on CAD, we evaluated the effects of TCV-116, an AT1-R antagonist, in a murine model of cardiac transplantation. METHODS AND RESULTS: Hearts of DBA/2 mice (H-2d) were transplanted heterotopically to B10.D2 mice (H-2d). Recipients were treated orally with TCV-116 (10 mg/kg per day), captopril (100 mg/kg per day), or vehicle only. Graft status, as assessed by palpation and inspection at laparotomy 70 days after transplantation, was preserved better in the TCV-116-treated group (P < .005) and in the captopril-treated group (P < .05) than in the vehicle-treated group. Intimal area in the graft coronary arterial wall decreased to 31% in the TCV-116-treated group (P < .001 versus vehicle-treated group) and to 34% (P < .005) in the captopril-treated group but was 45% in the vehicle-treated group. Fibrotic lesions of the left ventricle were less prominent in the TCV-116-treated (31%; P < .01 versus vehicle-treated group) and captopril-treated groups (33%; P < .05) than in the vehicle-treated group (54%). CONCLUSIONS: These findings show that AT1-R blockade is at least as effective as ACE inhibition in management of chronic allograft rejection and suggest that Ang II may play an important role in chronic allograft rejection.     

Mast cell-dependent neutrophil and mononuclear cell recruitment in immunoglobulin E-induced gastric reactions in mice

BACKGROUND & AIMS: Immunoglobulin (Ig) E-dependent reactions elicit an immediate response and can also result in a late-phase reaction that is characterized by the infiltration of leukocytes. This study assessed whether IgE-dependent late-phase responses can be elicited in the stomach wall of mice and examined the role of mast cells in this reaction. METHODS: IgE-dependent gastric inflammation was elicited in genetically mast cell-deficient KitW/KitW-v mice, the congenic normal (+/+) mice, and mast cell-deficient KitW/KitW-v mice that had undergone local and selective reconstitution of gastric mast cell populations. RESULTS: IgE-dependent gastric reactions were associated with mast cell degranulation and the infiltration of both neutrophils and mononuclear cells in normal mice, but no significant leukocyte infiltration was observed in mast cell-deficient KitW/KitW-v mice. By contrast, in mast cell-reconstituted KitW/KitW-v mice, IgE-dependent reactions were associated with the infiltration of neutrophils and mononuclear cells. CONCLUSIONS: These results show that late-phase reactions can occur during IgE-dependent gastric inflammation in the mouse and that the infiltration of both neutrophils and mononuclear cells that are observed during this reaction are mast cell dependent.     

The relationship between cardiac reactivity in the laboratory and in real life.

Objective: An excessive cardiovascular response to acute stress is a probable risk factor for cardiovascular (CV) disease. Such reactivity is usually assessed from the CV response to laboratory stressors. However, if it is a risk factor, correlated responses must occur in real life. Design: In the present study, we investigated the relationship between the heart rate (HR) response to five laboratory stressors and HR reactivity in the field. Measures: HR variation, the response to a real life stressor (public speaking), and the increase in HR with periods of self-reported tense arousal. Ambulatory HR, activity and posture were measured continuously over a 7-hr period. Results: The HR increase to laboratory stressors did not relate to HR variation consistently, but it did relate to the other two field measures. Conclusion: The results suggested that a tendency to increased HR reactivity may be a risk factor for cardiovascular disease when combined with exposure to stress. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Simvastatin suppresses glomerular cell proliferation and macrophage infiltration in rats with mesangial proliferative nephritis

Inhibition of 3-hydro-3-methylglutaryl coenzyme A reductase inhibits the production of mevalonate and has been shown to suppress proliferation in many cell types. Therefore, 3-hydro-3-methylglutaryl coenzyme A reductase inhibitors may have a beneficial effect in glomerular disease, because glomerular cell proliferation is a central feature in the active glomerular injury. This study examines the effect of simvastatin on glomerular pathology in a rat mesangial proliferative glomerulonephritis (GN) induced by anti-thymocyte antibody (anti-Thy 1.1 GN). There was no difference in the degree of the antibody and complement-mediated initial injuries between simvastatin-treated and control GN rats. The most pronounced feature of simvastatin-treated GN was the suppression of the early glomerular cell proliferation. The proliferative activity was maximal at day 4 after disease induction (26.5+/-7.0 of proliferating cell nuclear antigen-positive cells/glomerulus); however, approximately 70% of proliferation was suppressed by simvastatin treatment. At day 4 after disease induction, simvastatin administration also decreased alpha-smooth muscle actin expression in the glomerulus, which is a marker for mesangial cell activation. Inhibition of monocyte/macrophage recruitment into glomeruli by simvastatin was also a prominent feature. There was a 30% decrease in the number of glomerular ED-1+ cells by simvastatin treatment at day 2 after disease induction. Furthermore, simvastatin remarkably suppressed subsequent mesangial matrix expansion and type IV collagen accumulation in glomeruli. We also found that the platelet-derived growth factor expression was reduced in simvastatin-treated nephritic rats, which might simply reflect the reduction in mesangial cell proliferation and mesangial cellularity. There was no significant difference in plasma cholesterol or triglyceride levels between simvastatin- and vehicle-treated nephritic rats at day 2 and day 4, which corresponded to the times when simvastatin treatment resulted in a reduction in mesangial cell proliferation. In conclusion, this is the first report to find that mesangial cell proliferation and matrix expansion have been blocked by simvastatin in vivo. The protective effect of simvastatin in the matrix expansion in anti-Thy1.1 GN was partly by inhibition of mesangial cell proliferation and monocyte/ macrophage recruitment into glomeruli, which were independent of a change in circulating lipids.     

T cell differentiation and cytokine expression in late life

Elderly humans are at significant risk with regard to the incidence and severity of many infectious diseases and cancers. Current theory holds that these late-life vulnerabilities arise, in part, through age-related changes in immune function, particularly in the T lymphocyte lineage. Herein, we discuss how such factors as thymic involution and ongoing T cell differentiation in the peripheral tissues contribute to progressive and irreversible shifts in the state of differentiation of the mature T cell pool. We propose that, by late life, these processes yield a T cell compartment with a suboptimal balance of naive and memory T cell subsets, each with altered, subset-specific programs for cytokine gene expression. As such, the T cell compartment in late life may be more prone to immune deficiency or cytokine-mediated dysregulation in response to new or previously encountered pathogens.     

Chemokine and cell adhesion molecule mRNA expression and neutrophil infiltration in lipopolysaccharide-induced hepatitis in ethanol-fed rats

Neutrophil infiltration is a feature of alcoholic hepatitis (AH), and although the mechanism by which this occurs is unclear, it may involve a chemotactic gradient. We used lipopolysaccharide (LPS) to induce, in ethanol-fed rats, liver damage similar to that seen in AH. To our knowledge, this study is the first to examine the effect of ethanol on LPS-stimulated chemokine mRNA expression in this model. Hepatic cytokine-induced neutrophil chemoattractant (CINC)-1, CINC-2, monocyte chemoattractant protein-1 (MCP-1), macrophage inflammatory protein (MIP)-1beta, MIP-2, and eotaxin mRNA levels were elevated 1 to 3 hr post-LPS in both groups. Maximal expression of MIP-2 and MCP-1 mRNA was higher in ethanol-fed rats 1 hr post-LPS, whereas CINC-2 mRNA expression was elevated above controls at 12 to 24 hr. Hepatic intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 mRNA levels were elevated in both groups at 1 hr, whereas L-selectin expression in ethanol-fed rats was elevated above controls at 12 to 24 hr. Hepatic neutrophil infiltration was highest during maximal hepatocyte necrosis. These data suggest that cell adhesion molecules, in conjunction with elevated cytokines and the subsequently induced chemokines, may assist in the formation of a chemotactic gradient within the liver, causing the neutrophil infiltration seen both in this model and possibly in AH.     

Normoxic cardiopulmonary bypass reduces oxidative myocardial damage and nitric oxide during cardiac operations in the adult

The purpose of this study was to explain smoking habits amongst middle-aged men in Finland by describing their experiences of smoking and their attitudes towards smoking. As a pilot survey for a major health campaign targeted at 40-year-old men, the data for this study were collected using two questionnaires in connection with voluntary medical examinations. The first questionnaire was based on Prochaska's theory of stages of change in health behaviour. The second instrument was an attitude scale developed specifically for this study on the basis of Green and Kreuter's theory of factors influencing health behaviour. According to the results 31% of males aged 40 were regular smokers. Men with a lower level of education and out of work smoked more often than others. Non-smokers reported a better self-perceived health than smokers. Smoking cessation is a process in which men gradually proceed from one step to the next. In this study 12% of the men were in the contemplation stage and 11% in the preparation stage. One-quarter of the men had recently given up the habit and were in the action stage, while 2% had quit smoking over 6 months ago and were in the maintenance stage. One-quarter of the men regarded smoking as an integral part of their way of life and felt that public opinion towards smoking is hostile.     

Coronary angioplasty, bypass surgery, and retransplantation in cardiac transplant patients with graft coronary disease

BACKGROUND AND OBJECTIVE: Prior studies of laser tissue soldering (LTS) of epithelial skin have shown poor wound strength in the short-term; however, we hypothesize that greater tensile strength and healing properties will result from directing laser energy to the dermal aspect of the skin. The current study compares wound strength and histology in a rat skin flap model of epithelial and dermally applied LTS. STUDY DESIGN/MATERIALS AND METHODS: Skin flaps (2.5 x 4 cm) were raised and bisected on the dorsum of Sprague-Dawley rats. The center line of bisection was closed from a dermal approach by LTS (LTS-D, diode laser 15.9 W/cm2 + Columbia solder), the upper incision by epithelial LTS (LTS-E), and the lower incision by suturing (7-0 Vicryl). Wound skin strips (1-2 mm x 10 mm) were studied immediately (N = 14) and at 3 (N = 57), 7 (N = 31), and 10 (N = 28) days postoperatively and were subjected to tensiometric analysis. Histologic staining with hematoxylin and eosin and Mallory's trichrome methods were used to define wound architecture. RESULTS: No wound dehiscences were noted in any group. Greater immediate tensile strength was noted in wounds closed by LTS-D (521 +/- 61 g/cm2) versus LTS-E (342 +/- 65 g/cm2); however, this difference was not statistically significant (P = .08). By 3 days, both LTS-D (476 +/- 55 g/cm2) and LTS-E (205 +/- 37 g/cm2) maintained their initial strength; however, LTS-D and sutured (436 +/- 49 g/cm2) wounds were stronger (P < .05) than LTS-E. At 7 and 10 days, LTS-D (2,433 +/- 346 g/cm2 and 3,100 +/- 390 g/cm2) showed superior tensile strength (P < .05) compared to both LTS-E (1,542 +/- 128 g/cm2 and 2,081 +/- 219 g/cm2) and suturing (1,342 +/- 119 g/cm2 and 1,661 +/- 115 g/cm2). Histologic analysis of LTS-D wounds at 3 days showed full-thickness tissue apposition, complete epithelialization, and minimal inflammation or thermal injury. At 7 days, solder was present in the wounds. In contrast, LTS-E wounds at 3 days displayed lack of epithelialization secondary to thermal injury and partial-thickness tissue apposition. However by 7 days, epithelialization was complete with moderate scarring, and no solder was seen. Sutured samples appeared similar to LTS-D, except for poorer tissue apposition at the hypodermis. CONCLUSION: Our results show that skin flap wound healing after dermal LTS is superior to epithelial LTS and emphasizes the importance of site specificity in the utilization of this operative technique in reconstructive surgery.