Overcoming silos in health care systems through meso-level organisations - a case study of health reforms in New South Wales, Australia.

Fragmented care delivery is a barrier to improving health system performance worldwide. Investment in meso-level organisations is a potential strategy to improve health system integration, however, its effectiveness remains unclear. In this paper, we provide an overview of key international and Australian integrated care policies. We then describe Collaborative Commissioning - a novel health reform policy to integrate primary and hospital care sectors in New South Wales (NSW), Australia and provide a case study of a model focussed on older person's care. The policy is theorised to achieve greater integration through improved governance (local stakeholders identifying as part of one health system), service delivery (communities perceive new services as preferable to status quo) and incentives (efficiency gains are reinvested locally with progressively higher value care achieved). If effectively implemented at scale, Collaborative Commissioning has potential to improve health system performance in Australia and will be of relevance to similar reform initiatives in other countries.

Accuracy of reporting of Aboriginality on administrative health data collections using linked data in NSW, Australia.

BACKGROUND: Aboriginal people are under-reported on administrative health data in Australia. Various approaches have been used or proposed to improve reporting of Aboriginal people using linked records. This cross-sectional study used self-reported Aboriginality from the NSW Patient Survey Program (PSP) as a reference standard to assess the accuracy of reporting of Aboriginal people on NSW Admitted Patient (APDC) and Emergency Department Data Collections (EDDC), and compare the accuracy of selected approaches to enhance reporting Aboriginality using linked data. METHODS: Ten PSP surveys were linked to five administrative health data collections, including APDC, EDDC, perinatal, and birth and death registration records. Accuracy of reporting of Aboriginality was assessed using sensitivity, specificity, and positive and negative predictive values (PPVs and NPVs) and F score for the EDDC and APDC as baseline and four enhancement approaches using linked records: "Most recent linked record", "Ever reported as Aboriginal", and two approaches using a weight of evidence, "Enhanced Reporting of Aboriginality (ERA) algorithm" and "Multi-stage median (MSM)". RESULTS: There was substantial under-reporting of Aboriginality on APDC and EDDC records (sensitivities 84 and 77% respectively) with PPVs of 95% on both data collections. Overall, specificities and NPVs were above 98%. Of people who were reported as Aboriginal on the PSP, 16% were not reported as Aboriginal on any of their linked records. Record linkage approaches generally increased sensitivity, accompanied by decrease in PPV with little change in overall F score for the APDC and an increase in F score for the EDDC. The "ERA algorithm" and "MSM" approaches provided the best overall accuracy. CONCLUSIONS: Weight of evidence approaches are preferred when record linkage is used to improve reporting of Aboriginality on administrative health data collections. However, as a substantial number of Aboriginal people are not reported as Aboriginal on any of their linked records, improvements in reporting are incomplete and should be taken into account when interpreting results of any analyses. Enhancement of reporting of Aboriginality using record linkage should not replace efforts to improve recording of Aboriginal people at the point of data collection and addressing barriers to self-identification for Aboriginal people.

The impact of the mode of survey administration on estimates of daily smoking for mobile phone only users.

BACKGROUND: Over the past decade, there have been substantial changes in landline and mobile phone ownership, with a substantial increase in the proportion of mobile-only households. Estimates of daily smoking rates for the mobile phone only (MPO) population have been found to be substantially higher than the rest of the population and telephone surveys that use a dual sampling frame (landline and mobile phones) are now considered best practice. Smoking is seen as an undesirable behaviour; measuring such behaviours using an interviewer may lead to lower estimates when using telephone based surveys compared to self-administered approaches. This study aims to assess whether higher daily smoking estimates observed for the mobile phone only population can be explained by administrative features of surveys, after accounting for differences in the phone ownership population groups. METHODS: Data on New South Wales (NSW) residents aged 18 years or older from the NSW Population Health Survey (PHS), a telephone survey, and the National Drug Strategy Household Survey (NDSHS), a self-administered survey, were combined, with weights adjusted to match the 2013 population. Design-adjusted prevalence estimates and odds ratios were calculated using survey analysis procedures available in SAS 9.4. RESULTS: Both the PHS and NDSHS gave the same estimates for daily smoking (12%) and similar estimates for MPO users (20% and 18% respectively). Pooled data showed that daily smoking was 19% for MPO users, compared to 10% for dual phone owners, and 12% for landline phone only users. Prevalence estimates for MPO users across both surveys were consistently higher than other phone ownership groups. Differences in estimates for the MPO population compared to other phone ownership groups persisted even after adjustment for the mode of collection and demographic factors. CONCLUSIONS: Daily smoking rates were consistently higher for the mobile phone only population and this was not driven by the mode of survey collection. This supports the assertion that the use of a dual sampling frame addresses coverage issues that would otherwise be present in telephone surveys that only made use of a landline sampling frame.

Towards public health surveillance of intensive care services in NSW, Australia.

OBJECTIVES: Outbreaks of known and novel pathogens causing very severe illness increase the risk to public health in a globalised community and alarm the public. Intensive care units (ICUs) may be an underused setting for public health surveillance. This study investigates the electronic Record for Intensive Care (eRIC), an electronic clinical information and management system being developed for New South Wales ICUs, and its surveillance opportunity offerings. METHODS: The surveillance benefits being introduced by the eRIC were evaluated through consultation with stakeholders and the eRIC program team. The consultation process involved providing stakeholders with background information about the eRIC system. Based on the consultation, a draft data and information model for surveillance was developed. The model was evaluated using guidelines from the US Centers for Disease Control and Prevention. RESULTS: Population health stakeholders confirmed that the eRIC offers an appealing surveillance data source for pathogens and other hazards causing severe illness. Suggested application of the surveillance included, for known hazards, seasonal and pandemic influenza, enterovirus 71, Murray Valley encephalitis virus, enterohaemorrhagic Escherichia coli 0104:H4 and parechovirus. The proposed surveillance model uses syndromic rather than specific-cause surveillance. It may offer greater timeliness and sensitivity than relying on reporting of diagnoses of specific pathogens. Five syndromes derived from clinical pathways in the eRIC are proposed: severe acute respiratory disease, severe acute neurological disease, sepsis or septicaemia, jaundice or hepatitis, and acute renal failure. CONCLUSION: New intensive care clinical information systems offer a largely untapped resource for continuous, mainstream, rapid ICU surveillance of severe illness. A continuous, mainstream, rapid ICU surveillance facility that will readily adapt to emergency situations would be a valuable resource for protecting population health. This study establishes a firm basis on which ICU surveillance can be developed.

Tipranavir/Ritonavir (500/200 mg and 500/100 mg) Was Virologically Non-Inferior to Lopinavir/Ritonavir (400/100 mg) at Week 48 in Treatment-Naïve HIV-1-Infected Patients: A Randomized, Multinational, Multicenter Trial.

Ritonavir-boosted tipranavir (TPV/r) was evaluated as initial therapy in treatment-naïve HIV-1-infected patients because of its potency, unique resistance profile, and high genetic barrier. Trial 1182.33, an open-label, randomized trial, compared two TPV/r dose combinations versus ritonavir-boosted lopinavir (LPV/r). Eligible adults, who had no prior antiretroviral therapy were randomized to twice daily (BID) 500/100 mg TPV/r, 500/200 mg TPV/r, or 400/100 mg LPV/r. Each treatment group also received Tenofovir 300 mg + Lamivudine 300 mg QD. The primary endpoint was a confirmed viral load (VL) <50 copies/mL at week 48 without prior antiretroviral regimen changes. Primary analyses examined CD4-adjusted response rates for non-inferiority, using a 15% non-inferiority margin. At week 48, VL<50 copies/mL was 68.4%, 69.9%, and 72.4% in TPV/r100, TPV/r200, and LPV/r groups, respectively, and TPV/r groups showed non-inferiority to LPV/r. Discontinuation due to adverse events was higher in TPV/r100 (10.3%) and TPV/r200 (15.3%) recipients versus LPV/r (3.2%) recipients. The frequency of grade ≥3 transaminase elevations was higher in the TPV/r200 group than the other groups, leading to closure of this group. However, upon continued treatment or following re-introduction after treatment interruption, transaminase elevations returned to grade ≤2 in >65% of patients receiving either TPV/r200 or TPV/r100. The trial was subsequently discontinued; primary objectives were achieved and continuing TPV/r100 was less tolerable than standard of care for initial highly active antiretroviral therapy. All treatment groups had similar 48-week treatment responses. TPV/r100 and TPV/r200 regimens resulted in sustained treatment responses, which were non-inferior to LPV/r at 48 weeks. When compared with the LPV/r regimen and examined in the light of more current regimens, these TPV/r regimens do not appear to be the best options for treatment-naïve patients based on their safety profiles.

HLA alleles association with changes in bone mineral density in HIV-1-infected adults changing treatment to tenofovir-emtricitabine or abacavir-lamivudine.

BACKGROUND: There are limited data regarding the influence of human leukocyte antigen (HLA) polymorphisms on reduced bone mineral density (BMD). We investigated the relationship between HLA supertypes and BMD in HIV-infected adults changing their existing treatment to tenofovir-emtricitabine (TDF-FTC) or abacavir-lamivudine (ABC-3TC) in the STEAL study. METHODS: Lumbar spine and right hip BMD were measured by Dual-energy X-ray absorptiometry (DXA). HLA genotypes at the 2-digit level were classified into class I and II supertypes. Student's t-tests were used to test the association between HLA supertypes and changes in hip and spine BMD over 96 weeks for the whole cohort and stratified by randomised groups. The relationship between HLA supertypes and BMD was also assessed in the subgroup of participants that were naïve to both ABC and TDF at study entry. RESULTS: Class II supertypes were mainly associated with hip BMD change. Overall, compared to participants not carrying HLA-DQ3, participants expressing DQ3 had less bone loss over 96 weeks at both the hip and spine (hip: 0.003 vs. -0.006 g/cm2, 95%CI 0.002 to 0.017, p = 0.016; spine: 0.006 vs. -0.006 g/cm2, 95%CI 0.001 to 0.023, p = 0.041). In participants that were naïve to both ABC and TDF at baseline and randomised to TDF-FTC, DQ3 was significantly associated with less bone loss compared with those not carrying DQ3 (hip: 0.001 vs. -0.032 g/cm2; diff 0.033; 95%CI 0.017 to 0.049; p<0.001; spine: 0.007 vs. -0.023 g/cm2; diff 0.035; 95%CI 0.014 to 0.056; p = 0.001). CONCLUSIONS: In this cohort of HIV-infected adults, there was an association between bone status and HLA supertypes, particularly HLA-DQ3. TRIAL REGISTRATION: Clinicaltrials.gov NCT00192634.

Counselling improves follow-up HIV testing at Week 6 for HIV postexposure prophylaxis recipients.

Demographic and clinical variables of clients attending a sexual health centre in western Sydney from 1 July 2009 to 30 June 2011 were examined to determine if nonoccupational HIV postexposure prophylaxis (NPEP) was being dispensed according to national guidelines,(1) and to identify factors associated with completion of follow-up. The results showed that 95.8% of antiretroviral prescriptions were consistent with national guidelines.(1) Consultation with a social worker significantly improved attendance for six week follow up serology (P=0.027).

The influence of HLA supertype on thymidine analogue associated with low peripheral fat in HIV.

OBJECTIVES: To examine the relationship between human leukocyte antigen (HLA) genotype and body composition changes induced by thymidine analogue nucleoside reverse transcriptase inhibitor (NtRTI) use in HIV-positive individuals. DESIGN: Data collected during the Simplification with Tenofovir-Emtricitabine (TDF-FTC) or Abacavir-Lamivudine (ABC-3TC) (STEAL) study were analysed to examine the potential association of HLA genotypes with changes in body composition in treatment-experienced HIV-positive individuals. METHODS: Demographic, HIV-related, body composition and HLA genotyping data from the STEAL study were used in this analysis. The mean percentage peripheral fat at study baseline was compared in participants with and without prior NtRTI use. Analyses were also carried out for each HLA supertype strata, for five HLA genes, within the thymidine-exposed group. These comparisons were made using Mann-Whitney rank-sum tests. RESULTS: Participants with prior NtRTI use had a significantly lower baseline mean peripheral fat percentage compared to those without NtRTI use (31.9 vs. 34.7%; P = 0.0045). However, participants carrying one or more of the three particular HLA supertype alleles, A01, B08 and DQ2, showed no significant difference in mean peripheral fat percentage at baseline by NtRTI use. Among participants with prior NtRTI exposure, there were significant differences in mean peripheral fat by HLA A01, B08 and DQ2 allele expression compared to those without expression of these alleles (A01: 34.91% vs. no A01: 30.3%; P = 0.0087; B08: 36.2% vs. no B08: 31.1%; P = 0.0317; DQ2: 35.16% vs. no DQ2: 30.06%; P = 0.0081). CONCLUSION: This analysis suggests that HIV-infected individuals carrying HLA A01, B08 or DQ2 supertype alleles may be resistant to NtRTI-induced peripheral fat loss.

Switching virally suppressed, treatment-experienced patients to a raltegravir-containing regimen does not alter levels of HIV-1 DNA.

BACKGROUND: Current HIV-1 antiretroviral therapy (ART) greatly reduces virus replication but does not significantly affect the viral reservoir. Raltegravir, a recently introduced integrase inhibitor, could, at least theoretically, reduce residual viremia in patients on ART and affect the viral reservoir size. The aim of this study was to assess whether switching therapy in treatment-experienced patients that were virally suppressed to a raltegravir-containing regimen reduces the size of the viral reservoir, and if such treatment leads to a change in levels of HIV 2-LTR circles in this patient group. METHODS: 14 ART experienced individuals with a suppressed viral load (<50 HIV-1 RNA copies/mL plasma) at baseline (for at least 2 months) were switched to a raltegravir-containing regimen. Blood samples were taken at baseline and at ≥2 timepoints up to 48±6 weeks. Levels of total HIV-1 DNA and 2-LTR circles in peripheral blood mononuclear cells (PBMCs) were measured using real-time PCR assays. RESULTS: There was no significant change in HIV-1 total DNA levels over the study duration (p = 0.808), median slope 0.24 (conservative nonparametric 95% CI: -11.78, 26.23). Low levels of 2-LTR circles were detected in 2 patients. One had 16 copies/10(6) PBMCs at baseline and the other had 34 copies/10(6) PBMCs at week 51. CONCLUSIONS: The switch to a raltegravir containing regimen was not associated with a significant change in HIV-1 total DNA levels in this cohort. There were no observed changes in the levels of HIV-1 2-LTR circles associated with raltegravir treatment initiation.

Optimal antiretroviral therapy for aging.

The introduction of highly active antiretroviral therapy (HAART) has irrevocably changed the nature of the HIV epidemic in developed countries. Although the use of HAART does not completely restore health in HIV-infected individuals, it has dramatically reduced morbidity and mortality. Increases in life expectancy resulting from effective long-term treatment mean that the proportion of older people living with HIV has increased substantially in the past 15 years. Increasing age is associated with many complications including cardiovascular disease, neurological complications, kidney and liver dysfunction, and metabolic complications such as dyslipidaemia and diabetes. HIV infection and antiretroviral drugs have also been associated with similar complications to those seen with increasing age. The increase in HIV prevalence in older age groups has not been accompanied by the development of treatment guidelines or recommendations for appropriate antiretroviral therapy or clinical management in these patients.

Raltegravir and unboosted atazanavir dual therapy in virologically suppressed antiretroviral treatment-experienced HIV patients.

BACKGROUND: Because of the favourable safety and tolerability profiles of atazanavir (ATV) and raltegravir (RAL), attention has recently turned to the use of dual ATV plus RAL therapy as a nucleoside reverse transcriptase inhibitor-sparing treatment strategy in highly antiretroviral treatment (ART)-experienced HIV-infected patients. METHODS: A retrospective observational study was carried out to assess the maintenance of viral suppression and ART tolerability in 20 highly ART-experienced patients with viral suppression, who had been switched to RAL and unboosted ATV dual therapy, using data collected during standard-of-care visits. RESULTS: At 6, 12 and 18 months, viral load was maintained at <400 HIV RNA copies/ml, with only one participant recording a detectable viral load (150 copies/ml) at the 6-month time point. Stable CD4(+) T-cell counts were maintained throughout the study period. Five participants changed regimen during the 18-month follow-up, with the median time to switch being 9 months (range 2-12). In three cases, patients were changed from dual therapy because of adverse events while on the regimen. These included increased fatigue (two patients), persistently increased bilirubin (one patient) and gastrointestinal side effects (one patient). Two additional patients changed therapy: one patient added lamivudine and one ceased ATV to pre-empt a potential drug-drug interaction. All five patients who switched from ATV/RAL before 12 months follow-up maintained viral suppression, implying no disadvantage from switching to dual therapy. CONCLUSIONS: Dual therapy with ATV plus RAL maintained viral suppression in this small group of highly ART-experienced patients. Further investigation of this novel dual therapy regimen is warranted.

Clinical research in NSW: its role in HIV care and prevention.

Clinical research in NSW has contributed to some important breakthroughs in the understanding of many aspects for HIV transmission, pathogenesis and treatment. Researchers in NSW have played an important role in understanding the progression of HIV disease, the development and use of antiretroviral therapies and have continued to be involved in the understanding, management and prevention of HIV infection. National and international collaboration are essential in identifying and managing the complex factors required for the current management of HIV and the potential mechanisms for the future elimination of HIV.

Characterization of a Plasmodium falciparum macrophage-migration inhibitory factor homologue.

BACKGROUND: Macrophage-migration inhibitory factor (MIF), one of the first cytokines described, has a broad range of proinflammatory properties. The genome sequencing project of Plasmodium falciparum identified a parasite homologue of MIF. The protein is expressed during the asexual blood stages of the parasite life cycle that cause malarial disease. The identification of a parasite homologue of MIF raised the question of whether it affects monocyte function in a manner similar to its human counterpart. METHODS: Recombinant P. falciparum MIF (PfMIF) was generated and used in vitro to assess its influence on monocyte function. Antibodies generated against PfMIF were used to determine the expression profile and localization of the protein in blood-stage parasites. Antibody responses to PfMIF were determined in Kenyan children with acute malaria and in control subjects. RESULTS: PfMIF protein was expressed in asexual blood-stage parasites, localized to the Maurer's cleft. In vitro treatment of monocytes with PfMIF inhibited random migration and reduced the surface expression of Toll-like receptor (TLR) 2, TLR4, and CD86. CONCLUSIONS: These results indicate that PfMIF is released during blood-stage malaria and potentially modulates the function of monocytes during acute P. falciparum infection.

The frequency of BDCA3-positive dendritic cells is increased in the peripheral circulation of Kenyan children with severe malaria.

The ability of Plasmodium falciparum-infected erythrocytes to adhere to host endothelial cells via receptor molecules such as ICAM-1 and CD36 is considered a hallmark for the development of severe malaria syndromes. These molecules are also expressed on leukocytes such as dendritic cells. Dendritic cells are antigen-presenting cells that are crucial for the initiation of adaptive immune responses. In many human diseases, their frequency and function is perturbed. We analyzed the frequency of peripheral blood dendritic cell subsets and the plasma concentrations of interleukin-10 (IL-10) and IL-12 in Kenyan children with severe malaria and during convalescence and related these parameters to the adhesion phenotype of the acute parasite isolates. The frequency of CD1c(+) dendritic cells in children with acute malaria was comparable to that in healthy controls, but the frequency of BDCA3(+) dendritic cells was significantly increased. Analysis of the adhesion phenotypes of parasite isolates revealed that adhesion to ICAM-1 was associated with the frequency of peripheral blood CD1c(+) dendritic cells, whereas the adhesion of infected erythrocytes to CD36 correlated with high concentrations of IL-10 and low concentrations of IL-12 in plasma.

Frequencies of peripheral blood myeloid cells in healthy Kenyan children with alpha+ thalassemia and the sickle cell trait.

The high frequencies of both alpha+ thalassemia and the sickle cell trait (hemoglobin AS [HbAS]) found in many tropical populations are thought to reflect selection pressure from Plasmodium falciparum malaria. For HbAS, but not for alpha+ thalassemia, protection appears to be mediated by the enhanced phagocytic clearance of ring-infected erythrocytes. We have investigated the genotype-specific distributions of peripheral blood leukocyte populations in two groups of children living on the coast of Kenya: a group of healthy P. falciparum parasite-negative children sampled at cross-sectional survey during a period of low malaria transmission, and a group of children attending the hospital with acute malaria. We report distinctive distributions of peripheral blood myeloid dendritic cells and monocytes in children with alpha+ thalassemia and HbAS during healthy periods and disease, and suggest ways in which these might relate to the mechanisms of protection afforded by these conditions.

Increased fatty acid desaturation and enhanced expression of stearoyl coenzyme A desaturase protects pancreatic beta-cells from lipoapoptosis.

Increased availability of fatty acids causes cell death and dysfunction in beta-cell lines, isolated islets, and animal models of diabetes. From the MIN6 beta-cell line, we selected two subpools that are resistant to palmitate-induced apoptosis. Protection was not universal because palmitate-resistant cells remained sensitive to cytokine- and streptozotocin-induced apoptosis. Palmitate oxidation and incorporation into cholesterol ester (but not triglycerides) were significantly higher in palmitate-resistant cells than in control cells. Consistent with these findings, transcript profiling revealed increased expression in palmitate-resistant cells of several beta-oxidation genes as well as a 2.8-fold upregulation of stearoyl-CoA desaturase 1 (SCD1). Correspondingly, the oleate-to-palmitate ratio of palmitate-resistant cells was double that of palmitate-pretreated control cells. At least some of this additional oleate in palmitate-resistant cells was incorporated into cholesterol ester stored in the form of large cytosolic lipid bodies. However, blocking cholesterol ester formation did not render palmitate-resistant cells sensitive to palmitate-induced apoptosis. On the other hand, an inhibitor of SCD1, 10,12-conjugated linoleic acid, dose dependently overcame the resistance of palmitate-resistant cells to lipoapoptosis. Our results suggest that desaturation per se is more important in protecting beta-cells from the cytotoxic effects of palmitate than is the nature of neutral lipid storage pool thus generated.

Chronic effects of fatty acids on pancreatic beta-cell function: new insights from functional genomics.

Type 2 diabetes can be viewed as a failure of the pancreatic beta-cell to compensate for peripheral insulin resistance with enhanced insulin secretion. This failure is explained by both a relative loss of beta-cell mass as well as secretory defects that include enhanced basal secretion and a selective loss of sensitivity to glucose. These features are reproduced by chronic exposure of beta-cells to fatty acids (FAs), suggesting that hyperlipidemia might contribute to decompensation. Using MIN6 cells pretreated for 48 h with oleate or palmitate, we have previously defined alterations in global gene expression by transcript profiling and described additional secretory changes to those already established (Busch A-K, Cordery D, Denyer G, Biden TJ: Diabetes 51:977-987, 2002). In contrast to a modest decoupling of glucose-stimulated insulin secretion, FA pretreatment markedly enhanced the secretory response to an acute subsequent challenge with FAs. We propose that this apparent switch in sensitivity from glucose to FAs would be an appropriate response to hyperlipidemia in vivo and thus plays a positive role in beta-cell compensation for insulin resistance. Altered expression of dozens of genes could contribute to this switch, and allelic variations in any of these genes could (to varying degrees) impair beta-cell compensation and thus contribute to conditions ranging from impaired glucose tolerance to frank diabetes.

Host systemic and local nitric oxide levels do not correlate with rejection of pig proislet xenografts in mice.

The rejection of pig proislet xenografts in mice is a CD4 T cell-dependent process in which macrophages play an important role. To assess the potential for activated macrophages to act as effector cells in xenograft destruction, we have examined the relationship between proislet xenograft rejection, two principal markers of macrophage activation, transcription of inducible nitric oxide synthase (iNOS) and production of nitric oxide (NO), and their temporal relationship to intragraft cytokine gene expression. Xenograft rejection in CBA/H mice correlated with early induction of intragraft host iNOS mRNA and marked intragraft production of NO (reactive nitrogen intermediates, RNI). Intragraft mRNA expression for IFN-gamma, IL-1beta and TNF, cytokines associated with macrophage activation, was also found. These findings suggested that activated macrophages could be contributing to xenograft destruction via local NO-mediated toxicity at the graft site. To test the role of NO in this model: (1) Q-fever antigen (QFA) was administered to recipient mice in order to induce high systemic RNI levels and (2) in another experiment, pig proislets were transplanted into iNOS-/- mice. Treatment with QFA correlated with prolonged xenograft survival at 7 days post-transplant. Splenocytes from QFA-treated, but not control mice at 7 and 22 days post-transplant, exhibited inhibition of secondary xenogeneic mouse antiporcine mixed lymphocyte reaction (MLR) that was reversed by culture with the NOS inhibitor N-methylarginine (NMA). Despite continued elevated NO production, xenograft protection was temporary with complete rejection by day 22. Evidence that locally produced NO was not contributing to rejection was seen when pig proislets transplanted into iNOS-/- mice were rejected with normal kinetics; in these animals intragraft NO production was not detected (despite porcine iNOS gene expression). Failure of activated macrophages to achieve indefinite xenograft survival suggests that other factors are also required. Macrophage potential to effect either destructive or protective roles after pig proislet xenotransplantation suggests that such functions may depend on the site and magnitude of macrophage activation. Together these findings clearly demonstrate that high NO levels in the periphery are not damaging to xenogeneic islet tissue, neither host nor donor NO production is essential for islet xenograft rejection and consequently elevated plasma RNI levels do not represent a direct marker for rejection.