Adiponectin, leptin and insulin in breast milk: associations with maternal characteristics and infant body composition in the first year of life.

BACKGROUND/OBJECTIVES: Breastfeeding may protect against excessive weight gain during infancy. However, the breast milk components responsible for this effect are unknown. We examined the variation of three breast milk hormones (adiponectin, leptin and insulin) according to maternal characteristics and determined their association with infant body composition. SUBJECTS/METHODS: We studied a representative subset of 430 breastfed infants in the CHILD birth cohort. Breast milk was collected at 4 months postpartum and hormone concentrations were measured using the MesoScale Discovery System. Weight-for-length (WFL) and body mass index (BMI) z-scores were calculated according to the World Health Organization reference standard from infant anthropometrics measured at 4 months and 1 year. Maternal BMI and demographics were self-reported. RESULTS: Breast milk hormone concentrations varied widely between mothers. The geometric mean (range) was 19.4 (3.7-74.4) ngml-1 for adiponectin; 361 (31-3968) pgml-1 for leptin; and 589 (53-5557) pgml-1 for insulin. Maternal BMI was positively correlated with breast milk insulin (r=+0.40, P<0.0001) and leptin (r=+0.71, P<0.0001), but not adiponectin (r=-0.02, P=0.68). Breast milk hormone concentrations were also associated with maternal ethnicity, parity and breastfeeding exclusivity at sample collection. Independent of these factors and maternal diabetes, smoking and breastfeeding duration, higher breast milk leptin was associated with lower infant WFL z-score at 4 months (ß -0.67, 95% confidence interval (CI): -1.17, -0.17 for highest vs lowest quintile) and 1 year (ß -0.58, 95% CI: -1.02, -0.14). Insulin showed a U-shaped association, with intermediate concentrations predicting the lowest infant WFL z-score at 4 months (ß -0.51, 95% CI: -0.87, -0.15 for third vs lowest quintile) and 1 year (ß -0.35, 95% CI: -0.66, -0.04). Similar results were seen with infant BMI. Breast milk adiponectin was not significantly associated with infant body composition. CONCLUSIONS: Breast milk hormone concentrations were associated with several fixed and modifiable maternal characteristics. Higher concentrations of leptin and intermediate concentrations of insulin were associated with lower infant WFL in the first year of life.

Patterns of allergic sensitization and atopic dermatitis from 1 to 3 years: Effects on allergic diseases.

BACKGROUND: While allergic sensitization and atopic dermatitis (AD) are known to increase the risk for allergic diseases, the impact of different temporal and clinical patterns of sensitization and AD is less well defined. OBJECTIVE: We investigated patterns of sensitization and AD from early infancy to age 3, and the differential risk of developing allergic diseases within each pattern in a general cohort. METHODS: Children (n = 2629) from the Canadian Healthy Infant Longitudinal Development (CHILD) Study underwent skin prick tests and were assessed clinically for AD at ages 1 and 3 years. We applied an unsupervised latent class analysis (LCA) to the following 5 factors at these ages: AD, food sensitization, inhalant sensitization, poly-sensitization to foods and poly-sensitization to inhalants. The risks for developing asthma, allergic rhinitis and food allergy at 3 years were evaluated for each identified group. RESULTS: Five distinct classes were revealed by LCA: healthy (81.8%), atopic dermatitis (7.6%), inhalant sensitization (3.5%), transient sensitization (4.1%) and persistent sensitization (3.2%). Using healthy children as the baseline, children in the "atopic dermatitis" group had the next lowest risk for all allergic outcomes at 3 years; those in the "inhalant sensitization" group had the highest risk for allergic rhinitis; children in the "transient sensitization" group were at an increased risk for food allergy; while children in the "persistent sensitization" group had the highest risk for all allergic diseases. CONCLUSION AND CLINICAL RELEVANCE: There is substantial heterogeneity among allergen-sensitized children. Researchers and clinicians need to be aware of the non-specificity associated with labelling children simply as "atopic" and "non-atopic" without considering the timing of their atopic history, type of sensitization and AD status. Children with AD who were poly-sensitized to foods at an early age appear to be at greatest risk of developing other allergic diseases.

Pre-diagnostic genotyping identifies T1D subjects with impaired Treg IL-2 signaling and an elevated proportion of FOXP3+IL-17+ cells.

T-regulatory cells (Tregs) are essential for immune tolerance, and animal studies implicate their dysfunction in type 1 diabetes (T1D) pathogenesis. Tregs require interleukin-2 (IL-2) for their suppressive function, and variants in IL-2/IL-2R pathway genes have been associated with T1D. We previously reported that recent-onset T1D subjects have an increased population of FOXP3lo Tregs that secrete the pro-inflammatory cytokine, interleukin-17 (IL-17). We hypothesize that IL-2 signaling defects may drive T1D development by skewing protective Tregs towards an inflammatory Th17 phenotype. Overall, we found that the proportion of FOXP3+IL-17+ cells in T1D subjects pre-diagnosis was unchanged compared with healthy controls. However, stratification by IL2RA single-nucleotide polymorphisms revealed that T1D subjects with the rs3118470 CC risk variant have Tregs with IL-2 signaling defects and an increased proportion of FOXP3+IL-17+ cells before diagnosis. These data suggest a potential mechanism for genetically controlled loss of Treg function via dysfunctional IL-2 signaling in T1D.

Impact of maternal intrapartum antibiotics, method of birth and breastfeeding on gut microbiota during the first year of life: a prospective cohort study.

OBJECTIVE: Dysbiosis of the infant gut microbiota may have long-term health consequences. This study aimed to determine the impact of maternal intrapartum antibiotic prophylaxis (IAP) on infant gut microbiota, and to explore whether breastfeeding modifies these effects. DESIGN: Prospective pregnancy cohort of Canadian infants born in 2010-2012: the Canadian Healthy Infant Longitudinal Development (CHILD) Study. SETTING: General community. SAMPLE: Representative sub-sample of 198 healthy term infants from the CHILD Study. METHODS: Maternal IAP exposures and birth method were documented from hospital records and breastfeeding was reported by mothers. Infant gut microbiota was characterised by Illumina 16S rRNA sequencing of faecal samples at 3 and 12 months. MAIN OUTCOME MEASURES: Infant gut microbiota profiles. RESULTS: In this cohort, 21% of mothers received IAP for Group B Streptococcus prophylaxis or pre-labour rupture of membranes; another 23% received IAP for elective or emergency caesarean section (CS). Infant gut microbiota community structures at 3 months differed significantly with all IAP exposures, and differences persisted to 12 months for infants delivered by emergency CS. Taxon-specific composition also differed, with the genera Bacteroides and Parabacteroides under-represented, and Enterococcus and Clostridium over-represented at 3 months following maternal IAP. Microbiota differences were especially evident following IAP with emergency CS, with some changes (increased Clostridiales and decreased Bacteroidaceae) persisting to 12 months, particularly among non-breastfed infants. CONCLUSIONS: Intrapartum antibiotics in caesarean and vaginal delivery are associated with infant gut microbiota dysbiosis, and breastfeeding modifies some of these effects. Further research is warranted to explore the health consequences of these associations. TWEETABLE ABSTRACT: Maternal #antibiotics during childbirth alter the infant gut #microbiome.

The Crohn's disease-associated polymorphism in ATG16L1 (rs2241880) reduces SHIP gene expression and activity in human subjects.

Crohn's disease (CD) is a polygenic immune-mediated disease characterized by gastrointestinal inflammation. Mice deficient in the hematopoietic-restricted SH2 domain-containing inositolpolyphosphate 5'-phosphatase (SHIP) develop spontaneous CD-like ileal inflammation. Intriguingly, SHIP mRNA is not upregulated in biopsies from patients with ileal CD despite immune cell infiltration, but SHIP's role in human CD remains unknown. We analyzed SHIP mRNA expression and activity in biopsies and peripheral blood mononuclear cells (PBMCs) from control and treatment-naive subjects with ileal CD, and demonstrated that SHIP mRNA and activity were lower in hematopoietic cells in ileal biopsies and PBMCs from subjects with CD. In all tissues from our patient cohort and in PBMCs from a second healthy control cohort, subjects homozygous for the autophagy-related 16-like protein (ATG16L1) CD-associated gene variant (rs2241880), had low SHIP mRNA expression and activity. SHIP protein expression increased during autophagy and SHIP upregulation was dependent on ATG16L1 and/or autophagy, as well as the ATG16L1 CD-associated gene variant. Finally, homozygosity for the ATG16L1 risk variant and low SHIP mRNA expression is inversely related to increased (LPS+ATP)-induced IL-1ß production by PBMCs in our cohorts and was regulated by increased transcription of ILIB. These data suggest a novel mechanism by which the ATG16L1 CD-associated gene variant may predispose people to develop intestinal inflammation.

Infant gut microbiota and food sensitization: associations in the first year of life.

BACKGROUND: The gut microbiota is established during infancy and plays a fundamental role in shaping host immunity. Colonization patterns may influence the development of atopic disease, but existing evidence is limited and conflicting. OBJECTIVE: To explore associations of infant gut microbiota and food sensitization. METHODS: Food sensitization at 1 year was determined by skin prick testing in 166 infants from the population-based Canadian Healthy Infant Longitudinal Development (CHILD) study. Faecal samples were collected at 3 and 12 months, and microbiota was characterized by Illumina 16S rRNA sequencing. RESULTS: Twelve infants (7.2%) were sensitized to ≥ 1 common food allergen at 1 year. Enterobacteriaceae were overrepresented and Bacteroidaceae were underrepresented in the gut microbiota of food-sensitized infants at 3 months and 1 year, whereas lower microbiota richness was evident only at 3 months. Each quartile increase in richness at 3 months was associated with a 55% reduction in risk for food sensitization by 1 year (adjusted odds ratio 0.45, 95% confidence interval 0.23-0.87). Independently, each quartile increase in Enterobacteriaceae/Bacteroidaceae ratio was associated with a twofold increase in risk (2.02, 1.07-3.80). These associations were upheld in a sensitivity analysis among infants who were vaginally delivered, exclusively breastfed and unexposed to antibiotics. At 1 year, the Enterobacteriaceae/Bacteroidaceae ratio remained elevated among sensitized infants, who also tended to have decreased abundance of Ruminococcaceae. CONCLUSIONS AND CLINICAL RELEVANCE: Low gut microbiota richness and an elevated Enterobacteriaceae/Bacteroidaceae ratio in early infancy are associated with subsequent food sensitization, suggesting that early gut colonization may contribute to the development of atopic disease, including food allergy.

The Canadian healthy infant longitudinal development birth cohort study: biological samples and biobanking.

BACKGROUND: It is hypothesised that complex interactions between genetic and environmental factors give rise to allergy and asthma in childhood. The Canadian Healthy Infant Longitudinal Development (CHILD) study was designed to explore these factors. METHODS: CHILD is a longitudinal, general population birth cohort study following infants from mid-pregnancy to age 5 years. Over this time period, biological samples, questionnaires, clinical measures and environmental data are collected. RESULTS: A total of 3624 families have been recruited, and many thousands of samples and questionnaires have been collected, annotated, and archived. This report outlines the rationale and methodology for collecting and storing diverse biological samples from parents and children in this study, and the mechanisms for their release for analyses. CONCLUSIONS: The CHILD sample and data repository is a tremendous current and future resource and will provide a wealth of information not only informing studies of asthma and allergy, but also potentially in many other aspects of health relevant for Canadian infants and children.

Association of common single-nucleotide polymorphisms in innate immune genes with differences in TLR-induced cytokine production in neonates.

Significant variability in cytokine and chemokine expression after Toll-like receptor (TLR) stimulation has been observed between individuals. In this study, we determined the immunophenotypic variation in a cohort of 152 neonates associated with specific single-nucleotide polymorphisms (SNPs). We identified 23 SNPs in 12 genes of the innate immune system to be significantly associated with differential cytokine and chemokine production. SNPs in three gene families, namely STAT, IRF and SYK, accounted for most associations. These gene families are important signaling components of the innate anti-viral response. A potentially damaging non-synonymous SNP in the TLR3 gene (rs3775291) associated with significant differences in expression of interferon-γ after stimulation with the synthetic TLR3 ligand, poly (I:C). Additionally, a general increase in cytokine production was observed in subjects of Asian descent. This observation could be associated with differences in SNP genotype distribution between racial groups in our cohort. Taken together, our data suggest that particular aspects of the newborn innate response to TLR stimulation are closely associated with genetic variation. These findings provide the basis for detailed molecular dissection of cause-effect relationships between genotype and immune responses, and may account for inter-individual differences in response to vaccination and risk for infection and autoimmune disease.

HostSeq: a Canadian whole genome sequencing and clinical data resource.

HostSeq was launched in April 2020 as a national initiative to integrate whole genome sequencing data from 10,000 Canadians infected with SARS-CoV-2 with clinical information related to their disease experience. The mandate of HostSeq is to support the Canadian and international research communities in their efforts to understand the risk factors for disease and associated health outcomes and support the development of interventions such as vaccines and therapeutics. HostSeq is a collaboration among 13 independent epidemiological studies of SARS-CoV-2 across five provinces in Canada. Aggregated data collected by HostSeq are made available to the public through two data portals: a phenotype portal showing summaries of major variables and their distributions, and a variant search portal enabling queries in a genomic region. Individual-level data is available to the global research community for health research through a Data Access Agreement and Data Access Compliance Office approval. Here we provide an overview of the collective project design along with summary level information for HostSeq. We highlight several statistical considerations for researchers using the HostSeq platform regarding data aggregation, sampling mechanism, covariate adjustment, and X chromosome analysis. In addition to serving as a rich data source, the diversity of study designs, sample sizes, and research objectives among the participating studies provides unique opportunities for the research community.

Toll-like receptors 2 and 4 and the cryopyrin inflammasome in normal pregnancy and pre-eclampsia.

OBJECTIVE: Pre-eclampsia involves a maternal inflammatory response that differs from both normal pregnancy and normotensive intrauterine growth restriction (IUGR). Our objective was to examine neutrophil Toll-like receptor (TLR), cryopyrin, nuclear factor-kappaB (NF-kappaB) subunit and interleukin-1beta (IL-1beta), and inflammatory cytokine profiles in women with pre-eclampsia or normotensive IUGR, as well as in normal pregnancy and non-pregnancy controls. DESIGN AND METHOD: A case-control study was performed. We examined the messenger RNA (mRNA) and protein expressions of TLR4 and TLR2, mRNA levels of cryopyrin, IL-1beta, NF-kappaB subunits p50 and p65, as well as maternal serum inflammatory cytokine profiles (IL-2, IL-6, tumour necrosis factor-alpha [TNF-alpha], interferon-gamma [IFN-gamma] and IL-10) in women with and without pre-eclampsia using real-time reverse transcription polymerase chain reactions, flow cytometry and multiplex immunoassays. SETTING: A single tertiary maternity hospital in Vancouver, Canada. POPULATION: Women with early-onset pre-eclampsia (<34 weeks of gestation, n = 25), women with late-onset pre-eclampsia (>or=34(+0) weeks of gestation, n = 25), women with normotensive IUGR (n = 25), women with normal pregnancy (n = 75) and non-pregnancy (n = 25) controls. RESULTS: Women with pre-eclampsia (as a single combined group of early- and late-onset, and particularly in women with early-onset pre-eclampsia) had increased TLR2 and TLR4 mRNA and protein expressions elevated cryopyrin, NF-kappaB subunit, and IL-1beta mRNA expression, and TNF-alpha:IL-10 and IL-6:IL-10 ratios compared with other groups. CONCLUSIONS: These data suggest that TLRs and cryopyrin may modulate the innate immune response of the maternal syndrome of pre-eclampsia, and might also trigger the differential inflammatory response existing between early onset pre-eclampsia and normotensive IUGR.

Primary immunodeficiency diseases: a practical guide for clinicians.

Primary immunodeficiency diseases (PIDs) are genetically determined disorders of the immune system resulting in greatly enhanced susceptibility to infectious disease, autoimmunity and malignancy. While individual PIDs are rare, as a group, it is estimated that between 1:2000 and 1:10 000 live births are affected by a PID. Moreover, PIDs can present at any age from birth to adulthood, posing a considerable challenge for the practising physician to know when and how to work-up a patient for a possible immune defect. In this review, we outline the basic organisation of the human immune system and the types of infections that occur when elements of the immune system are dysfunctional. Importantly, we provide practical guidelines for identifying patients who should be referred for assessment of possible immunodeficiency and an overview of screening investigations and effective therapeutic options available for these patients.

Impact of maternal pre-pregnancy overweight on infant overweight at 1 year of age: associations and sex-specific differences.

BACKGROUND: Maternal overweight or obesity (OWOB) is linked to gestational diabetes, fetal macrosomia and higher rates of caesarean delivery. OBJECTIVES: The study aims to assess whether maternal pre-pregnancy OWOB is associated with infant overweight in a sex-dependent manner, independent of microbiota-altering variables. METHODS: Weight and length measurements of 955 mother-infant pairs were obtained from the Canadian Healthy Infant Longitudinal Development cohort. Maternal pre-pregnancy weight was defined as follows: normal, overweight (25 ≤ body mass index < 30) and obese (body mass index ≥ 30). Age and sex-adjusted weight-for-length z-scores >97th percentile were classified as infant overweight at age 1 year. Associations between pre-pregnancy and infant overweight were determined by linear and logistic regression, adjusting for covariates. RESULTS: Maternal pre-pregnancy OWOB were associated with infant weight-for-length and overweight risk at 1 year. Except for pre-pregnancy obesity, these associations were not attenuated appreciably after adjustment for birth mode, exclusivity of breastfeeding, exposure to antibiotics and infant sex. Yet only boys born to mothers with obesity were three times more likely to become overweight at age 1 independent of microbiota-altering variables. Pre-pregnancy obesity was associated with weight-for-length in male and female infants. CONCLUSIONS: Maternal pre-pregnancy OWOB increases the risk of infant overweight, and this association is more evident in male infants.

Infant gut immunity: a preliminary study of IgA associations with breastfeeding.

Secretory immunoglobulin A (IgA) plays a critical role in gut mucosal immune defense. Initially provided by breastmilk, IgA production by the infant gut is gradually stimulated by developing gut microbiota. This study reports associations between infant fecal IgA concentrations 4 months after birth, breastfeeding status and other pre/postnatal exposures in 47 infants in the Canadian Healthy Infant Longitudinal Development cohort. Breastfed infants and first-born infants had higher median fecal IgA concentrations (23.11 v. 9.34 µg/g protein, P<0.01 and 22.19 v. 8.23 µg/g protein, P=0.04). IgA levels increased successively with exclusivity of breastfeeding (ß-coefficient, 0.37, P<0.05). This statistical association was independent of maternal parity and household pets. In the absence of breastfeeding, female sex and pet exposure elevated fecal IgA to levels found in breastfed infants. In addition to breastfeeding, infant fecal IgA associations with pre/postnatal exposures may affect gut immunity and risk of allergic disease.

Changes in myoplasmic sodium concentration during exposure to lactate in perfused rat heart.

OBJECTIVE: The free myoplasmic sodium concentration ([Na+]i) is thought to change during acidosis and ischaemia in the mammalian heart. Although there are methods available for measuring [Na+]i in isolated cells or small preparations, none is very effective in the isolated heart, which is one of the best models for studying ischaemia. The use of a fluorescent sodium indicator, sodium binding benzofuran isophthalate (SBFI), to measure [Na+]i in the isolated, perfused rat heart is described. This method was used to estimate the activity of the Na/H exchanger under conditions resembling some aspects of ischaemia. METHODS: The acetoxymethyl ester of SBFI was loaded into the perfused rat heart via the perfusate. Fluorescence signals developed on loading and were shown to be [Na+]i sensitive. An in vivo calibration technique allowed the fluorescence signals to be converted to [Na+]i. RESULTS: The [Na+]i under control conditions was 6.4 mM. When the hearts were exposed to 20 mM lactate at the normal extracellular pH (pHo = 7.4) for 10 min, developed pressure fell abruptly and then slowly recovered [Na+]i rose slowly by 1.5 mM during lactate exposure and paralleled the recovery of pressure. Exposure to lactate at reduced extracellular pH (pHo = 6.4) produced a larger rise in [Na+]i. In the presence of the Na/H exchange inhibitor 5-(N-methyl-N-isobutyl)-amiloride exposure to lactate caused a small fall in [Na+]i at pHo 7.4 and 6.4. CONCLUSIONS: The advantages and disadvantages of SBFI as an [Na+]i indicator in the perfused heart are described. The Na/H exchanger is an important regulator of [Na+]i during lactate induced intracellular acidosis, and it remains active in the presence of a substantial extracellular acidosis. This shows that low pHo does not necessarily inhibit the Na/H exchanger. The use of SBFI to estimate [Na+]i in the perfused heart is novel and has considerable potential for studies of ischaemia and related conditions.

Mechanisms of tolerance induction after intrathymic islet injection: determination of the fate of alloreactive thymocytes.

BACKGROUND: Intrathymic (IT) administration of antigen when combined with peripheral T-cell depletion has been shown to induce operational tolerance in a wide range of experimental protocols. IT injection of pancreatic islets has been demonstrated not only to induce tolerance to alloantigen but also to prevent the development of autoimmune beta-cell destruction in models of type I diabetes. However, little is known about the mechanisms involved in tolerance induction after IT islet injection. METHODS AND RESULTS: A protocol for the induction of tolerance to fully allogeneic (C57BL/10; H2b) peripheral islet allografts was developed in CBA/Ca (H2k) recipients by the IT injection of allogeneic islets combined with depletion of peripheral CD4+ T cells. This protocol was based upon our own data and those of others showing that CD4+ T cells play a critical role in islet allograft rejection. Using this regimen, donor-type peripheral islet allografts survived indefinitely whereas third-party grafts were rejected. To determine the fate of alloreactive thymocytes that recognize donor major histocompatibility complex antigens via the direct pathway, T-cell receptor transgenic mice specific for the major histocompatibility complex class I molecule Kb (BM3 and DES) were used as recipients. IT injection of islets expressing the specific alloantigen Kb resulted in clonal deletion of alloreactive thymocytes in T-cell receptor transgenic recipients. No evidence of clonal inactivation in the residual peripheral alloreactive population was observed in this system. CONCLUSIONS: IT injection of allogeneic islets and concomitant CD4+ T-cell depletion is able to induce donor-specific unresponsiveness. One mechanism responsible for this unresponsiveness is the clonal deletion of thymocytes that recognize alloantigen via the direct pathway.

Islet allograft rejection can be mediated by CD4+, alloantigen experienced, direct pathway T cells of TH1 and TH2 cytokine phenotype.

BACKGROUND: It is widely believed that Thl cells that secrete interferon-gamma are primarily involved in the rejection of allografts whereas Th2 cells [interleukin(IL) 4 and IL-10] are thought to be protective of this process. However, the exact role and specificity of these helper T lymphocytes in mediating allograft damage is presently unknown. METHODS: Th0, Th1, and Th2 cell lines specific for the class II MHC molecule H2IAb were adoptively transferred into T cell deficient, syngeneic, diabetic mice before transplantation of fully allogeneic C57BL/10 (H2b) or (CBKxBALB/c)F1 (H2k/d+Kb) islet grafts. T cells were 5-(and-6-)-carboxyfluorescein diacetate succinimidyl ester- (CFSE) labeled to allow detection, immunohistochemistry was performed, and IL-4 transcripts within the rejected islet grafts were quantified by reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: Adoptive transfer (IV) of Th0-, Th1-, and Th2 IAb-specific T cells resulted in rejection of H2b islet allografts. CFSE-labeling demonstrated that these T cells were able to home to the graft site. CD4+ T cells and CD11b+ macrophages were present within the graft after adoptive transfer of both Thl and Th2 cells. Interestingly, CD8+ T cells and B cells were absent from these rejecting grafts. Even when Th2 cells were introduced directly at the graft site, prompt rejection was still observed despite the presence of increased IL-4 mRNA expression within the islet allografts. CONCLUSIONS: Th2 and Th0 alloreactive CD4+ T helper cells can reject islet grafts with similar efficiency to Th1 cells. These results suggest that deviation of the immune response from a Th1 to Th2 phenotype will not be sufficient to allow successful engraftment of allogeneic organs or tissues.

Fas ligand-transfected myoblasts and islet cell transplantation.

BACKGROUND: Expression of Fas ligand (FasL, CD95L) within the local environment of an allograft may protect from rejection by inducing apoptosis of infiltrating T cells. However, there is mounting evidence that ectopic expression of FasL stimulates an inflammatory response and targets the FasL-expressing tissue for destruction. Given the potential therapeutic applicability of FasL-based immune protection, we sought to determine whether ectopic FasL expression was detrimental and to analyze the inflammatory response induced by ectopic FasL expression in the absence of any confounding allo-immune responses. METHODS AND RESULTS: Two myoblast cell lines expressing different levels of functional FasL were produced. Co-implantation of FasL-expressing myoblasts with syngeneic islets allowed examination of the inflammatory response induced by ectopic FasL expression. In contrast to the suggested benefits of localized FasL expression, islets co-implanted with FasL-expressing myoblasts were destroyed in a vigorous inflammatory response predominated by neutrophils. Interestingly, FasL expression also had a marked anti-tumor effect. CONCLUSIONS: Unless FasL-dependent neutrophil-mediated inflammation can be prevented, it is unlikely that this strategy will be useful for preventing allograft rejection.

The visualization of T cell responses.

Transplanting allogeneic grafts is still significantly hampered by the rejection process, despite the use of powerful immunosuppressive agents. The T cell is recognized as playing a central role in the process of rejection, and it is believed that graft tolerance will ultimately be achieved by immunological manipulation of this cell (1, 2). As immunologists strive to define the role of the T cell in the fundamental processes of immunity and tolerance, new methods are emerging that will facilitate visualization of the T cells directly involved in the rejection response (3, 4). This overview addresses the visualization of T cell responses as made possible by these technological developments.

Intracellular calcium and myocardial function during ischemia.

Cardiac ischemia causes a rapid decline in mechanical performance and, if prolonged, myocardial cell death occurs on reperfusion. The early decline in mechanical performance could, in principle, be caused either by reduced intracellular calcium release or by reduced responsiveness of the myofibrillar proteins to calcium. It is now known that intracellular calcium rises during ischemia and that the early decline in mechanical performance is caused largely by the inhibitory effects of phosphate and protons on the myofibrillar proteins. The rise of intracellular calcium during ischemia is related to the acidosis and is probably caused by calcium influx on the Na/Ca exchanger. This is triggered by a rise in intracellular sodium which enter the cell in exchange for protons on the Na/H exchanger. Intracellular calcium rises still further on reperfusion the elevation of calcium and the degree of muscle damage are closely correlated.