Serum interleukin-4 is elevated in clinical drug-induced liver injury.

BACKGROUND: Drug-induced liver injury (DILI) remains a challenging diagnosis due to an absence of specific biomarkers. DILI due to volatile anaesthetics (VA-DILI) is characterised by trifluoroacetyl and CYP2E1 antibodies, but may not be seen for weeks after injury. Interleukin-4 (IL-4) may be involved in the production of these antibodies and may serve as a clinically useful early biomarker of VA-DILI. AIM: To prospectively compare serum IL-4 levels between patients who develop VA-DILI and controls following exposure to the volatile anaesthetic. METHODS: A nested case-control study of patients exposed to VA during surgery was conducted. Thirteen DILI cases were identified from the original cohort, and 26 controls were matched according to age, sex and VA agent. Serum samples were collected before and 48-96 h after VA exposure, and analysed for IL-4 using quantitative enzyme-linked immunosorbent assay techniques. RESULTS: There was a statistically significant difference in serum IL-4 in post-VA samples between DILI cases and controls (control: 0.030 pg/mL, IQR: 0.030 - 0.030 pg/mL vs DILI: 0.044 pg/mL, IQR: 0.030 - 0.061 pg/mL; p = 0.039). A greater proportion of DILI cases had post-VA IL-4 levels above the assay lower limit of detection compared to controls (control: 23% vs DILI: 69%; p = 0.013). CONCLUSION: IL-4 is a potential biomarker of DILI. Clinical diagnosis and understanding of DILI disease mechanisms may be improved by further investigation of novel biomarkers, and this IL-4 signal in serum is important as proof of concept for prospective study designs.

Intravenous thrombolysis for multi-ethnic Asians with acute ischaemia stroke in Malaysian public primary stroke centres versus acute stroke ready hospitals: Comparison of real-world clinical outcomes.

INTRODUCTION: Intravenous thrombolysis (IVT) with recombinant tissue plasminogen activator is beneficial in acute ischaemic stroke (AIS). We aim to compare the realworld clinical outcomes and service efficiency of IVT in Malaysian primary stroke centres (PSCs) versus acute stroke ready hospitals (ASRHs). MATERIALS AND METHODS: We conducted a multi-centre cohort study involving 5 PSCs and 7 ASRHs in Malaysia. Through review of medical records of AIS patients who received IVT from 01 January 2014 to 30 June 2021, real-world data was extracted for analysis. Univariate and multivariate regression models were employed to evaluate the role of PSCs versus ASRHs in post-IVT outcomes and complications. Statistical significance was set at p<0.05. RESULTS: A total of 313 multi-ethnic Asians, namely 231 from PSCs and 82 from ASRHs, were included. Both groups were comparable in baseline demographic, clinical, and stroke characteristics. The efficiency of IVT delivery (door-toneedle time), functional outcomes (mRS at 3 months post- IVT), and rates of adverse events (intracranial haemorrhages and mortality) following IVT were comparable between the 2 groups. Notably, 46.8% and 48.8% of patients in PSCs and ASRHs group respectively (p=0.752) achieved favourable functional outcome (mRS≤1 at 3 months post-IVT). Regression analyses demonstrated that post-IVT functional outcomes and adverse events were independent of the role of PSCs or ASRHs. CONCLUSION: Our study provides real-world evidence which suggests that IVT can be equally safe, effective, and efficiently delivered in ASRHs. This may encourage the establishment of more ASRHs to extend the benefits of IVT to a greater proportion of stroke populations and enhance the regional stroke care.

Real-world analysis of clinical and economic impact of 21-gene recurrence score (RS) testing in early-stage breast cancer (ESBC) in Ireland.

PURPOSE: Results from TAILOR-X suggest that up to 70% of hormone receptor-positive (HR+) node-negative (N0) ESBC patients (pts) may avoid chemotherapy (CT) with RS ≤ 25. We assess clinical and economic impacts of RS testing on treatment using real-world data. METHODS: From October 2011 to February 2019, a retrospective, cross-sectional observational study was conducted of HR+ N0 ESBC pts who had RS testing in Ireland. Pts were classified low risk (RS ≤ 25) and high risk (RS > 25). Clinical risk was calculated. Data were collected via electronic patient records. Cost data were supplied by the National Healthcare Pricing Regulatory Authority. RESULTS: 963 pts. Mean age is 56 years. Mean tumour size is 1.7 cm. 114 (11.8%), 635 (66%), 211 (22%), 3 (0.2%) pts had G1, G2, G3 and unknown G, respectively. 796 pts (82.8%) low RS, 159 (16.5%) high RS and 8 pts (0.7%) unknown RS. 263 pts (26%) were aged ≤ 50 at diagnosis; 117 (45%) had RS 0-15, 63 (24.5%) 16-20, 39 (15.3%) 21-25 and 40 (15.2%) RS 26-100. 4 pts (1.5%) had unknown RS. Post-RS testing, 602 pts (62.5%) had a change in CT decision; 593 changed to hormone therapy (HT) alone. In total, 262 pts received CT. Of pts receiving CT; 138 (53%) had RS > 25, 124 (47%) had RS ≤ 25. Of pts aged ≤ 50, 153 (58%) had high clinical risk, of whom 28 had RS 16-20. Assay use achieved a 62.5% change in treatment with 73% of pts avoiding CT. This resulted in savings of €4 million in treatment costs. Deducting assay costs, savings of €1.9 million were achieved. CONCLUSION: Over the 8 years of the study, a 62.5% reduction in CT use was achieved with savings of over €1,900,000.

Alendronate use in older patients with reduced renal function: challenges and opportunities in clinical practice.

In this retrospective cohort study, alendronate use among older osteoporosis patients (age>65 years) with reduced renal function (creatinine clearance<35ml/min) was not associated with significant deterioration in renal function from baseline nor increased incidence of osteoporotic fractures or acute kidney injury, compared with patients conservatively managed with only calcium/vitamin D supplementation. INTRODUCTION: Oral bisphosphonates are not recommended in patients with creatinine clearance (CrCl) <35ml/min, although this is not supported by post hoc analyses of pivotal oral bisphosphonate studies. As both osteoporosis and renal insufficiency are more prevalent with advancing age, it is important to determine the safety and efficacy of oral bisphosphonates among these patients. METHODS: Patients with CrCl <35ml/min on alendronate (group A, n=98), with CrCl <35ml/min conservatively managed (group B, n=96), and with CrCl ≥35ml/min on alendronate (group C, n=96) were followed up to 22 months. Primary outcomes were mean change in CrCl from baseline in group A compared with groups B and C, respectively. Secondary outcomes were the incidence of osteoporotic fractures and adverse events between groups. RESULTS: There was no significant change in CrCl from baseline when comparing group A (-1.53±6.83ml/min) with group B (0.59±5.17ml/min) (p=0.075), and group A with group C (-3.71±7.54ml/min) (p=0.163). There was no significant increase in incidences of osteoporotic fractures in group A compared with group B (adjusted relative risk (aRR) 2.02, 95% confidence interval (CI) 0.64-6.37) and group A compared with group C (aRR 1.15, 95% CI 0.46-2.89). There was no significant difference in incidences of acute kidney injury (AKI) in group A compared with group B (aRR 0.48, 95% CI 0.20-1.12). Although statistically non-significant, there was an increase in AKI incidence in group A compared with group C (RR 7.84, 95% CI 0.98-62.66). CONCLUSION: Among patients with CrCl <35ml/min, alendronate therapy was not associated with significant deterioration in renal function from baseline. Although not powered for secondary outcomes, there were no statistically significant differences in osteoporotic fracture or AKI incidence between the groups.

Clinical outcomes of acute stroke thrombolysis in neurologist and non-neurologist centres - A comparative study in Malaysia.

BACKGROUND: Thrombolytic therapy with intravenous alteplase is a well-established treatment for acute ischaemic stroke (AIS). However, in Malaysia, treatment prescription is often limited by the availability of neurologists. The objective was to compare the outcomes of acute stroke thrombolysis therapy prescribed by neurologists in the Seberang Jaya Hospital (SJH) and non-neurologists in the Taiping Hospital (TH). METHODS: In this cross-sectional study, all AIS patients who received thrombolytic therapy in SJH and TH between January 2012 and September 2019 were included. Clinical data was extracted from admission records. The outcomes assessed were the percentage of patients who achieved excellent functional outcome at 3 months (modified Rankin scale of 0 to 1), rates of symptomatic intracranial haemorrhage (SICH), and mortality. RESULTS: A total of 63 AIS patients who received thrombolytic therapy were included, of which 37 patients (58.7%) were treated in SJH. The median NIHSS on admission was 12 in SJH and 11.5 in TH. In all 21.6% of patients from SJH and 30.7% of patients from TH achieved favourable functional outcome at 3 months (p=0.412). There were no significant differences between the two centres in terms of the rates of SICH (10.8% in SJH and 3.8% in TH, p=0.314) and 3-month mortality (24.3% versus 12.5%, p=0.203). CONCLUSION: The 3-month functional outcomes and complication rates of stroke thrombolysis in hospitals with or without neurologists are not significantly different. Thus non-neurologist hospitals may be able to provide thrombolysis service to AIS patients safely and effectively.

Atypical meningioma with nodal metastasis: A case report.

INTRODUCTION: Meningiomas are common and usually benign central nervous system neoplasms. These neoplasms are graded into three groups which differ in biological behaviour. Atypical meningioma is an intermediate grade (Grade 2) tumour that is rarely associated with metastases compared to higher grade (Grade 3) meningiomas. CASE REPORT: A 68-year-old lady with a history of multiple craniotomies and hemifacial resections for meningothelial meningioma currently underwent orbital exenteration, tumour debulking and cervical nodal excision for tumour recurrence. Histopathological examination of the tumour showed atypical meningioma, with cervical nodal metastasis. DISCUSSION: This case report presents a rare finding of lymph node metastasis associated with atypical meningioma. The previous history of surgical resection is a known risk factor for metastasis for low to intermediate grade meningioma. Tumour biology and histology are predictors of metastasis. Haematogenous dissemination is the commonest route of metastasis. No standardised management protocol has been developed and the prognosis remains unknown.

Leptomeningeal Relapse of Embryonal Rhabdomyosarcoma after 15 years.

Aim Rhabdomyosarcoma (RMS) is the most common malignant soft tissue tumour of childhood. We present the case of a late relapse of RMS to the leptomeninges after 15 years. Methods A 20 year old male presented with a 3 week history of headaches and nausea. He previously had RMS of his right ear diagnosed at age 5 years which was treated with concurrent chemoradiotherapy. An MRI Brain and Spine confirmed extensive leptomeningeal disease and CSF analysis confirmed the presence of recurrent embryonal RMS. Results He completed two cycles of cyclophosphamide and topotecan followed by 45Gy/25Fr of craniospinal radiotherapy. Conclusion Late relapses beyond five years can be seen in up to 9% of patients, however very late recurrences (>10 years) are exceedingly rare. Molecular based methods such as gene expression profiling can aid risk stratification and survivorship clinics may become increasingly useful in following patients with high risk features.

The efficacy of intratympanic steroid injection for the treatment of idiopathic sudden sensorineural hearing loss.

OBJECTIVE: A systemic steroid is known to have a potential to recover hearing after idiopathic sudden sensorineural hearing loss (ISSHL). However, lately many centres have introduced the use of intratympanic (IT) steroid therapy as an option. We reviewed our experience in the treatment of patients with ISSHL looking at the overall success of using systemic steroid, IT steroid injections as salvage therapy and primary IT steroid injection. METHODS: A retrospective study was conducted on 20 patients who had suffered from ISSHL from January 1, 2012, to December 31, 2017. ISSHL is defined as a rapid decline in hearing over three days or less affecting three or more frequencies by 30dB or greater. Comparison between the mode of steroid therapies and improvement in patients was done. At least 15dB improvement in pure tone audiogram (PTA) was considered as successful therapeutic intervention. RESULTS: Twenty male and female patients who fit the inclusion and exclusion criteria were included. The mean age of the patients was 41.4 years with a range from 13 to 72 years. Ninety percent patients presented with unilateral ISSHL involvement. Eight ears of patients who received systemic steroid therapy alone had improved hearings (75%). Out of seven ears from six patients who received salvage therapy, four ears (57.1 %) had improvement in PTA. Seven ears showed improvement in PTA from a total of eight patients who primarily received IT injections. CONCLUSIONS: IT steroid therapy promises a favourable outcome in the improvement of the hearing, as compared to systemic steroid administration. Its usage is recommended not only for salvage therapy but should be used as primary treatment especially in those with co- morbidities.

Cheiroarthropathy and tendinopathy in diabetes.

Joint problems commonly occur in people with diabetes. Cheiroarthropathy affects the hands and results in painless limited finger joint extension, appearing to be associated with longer diabetes duration and the presence of microvascular complications. The prevalence of cheiroarthropathy seems to be falling, perhaps as a result of improvements in glycaemic management. Non-enzymatic glycation of collagen results in abnormally crosslinked protein resistant to degradation with subsequent increased build-up of collagen in joints. The management of cheiroarthropathy is predominantly conservative, with occupational and hand therapy at the forefront. Tendinopathy is more common in people with diabetes than those without, and is associated with obesity and insulin resistance. As with cheiroarthropathy, the exact causative mechanism of tendinopathy in diabetes is not known, but may be linked to inflammation, apoptosis and increased vascularity of affected tendons, driven by hyperinsulinaemia. Local fat pads have also been suggested to play a role in the pathogenesis of tendinopathy.

International cancer seminars: a focus on kidney cancer.

Recent years have seen important advances in our understanding of the etiology, biology and genetics of kidney cancer. To summarize important achievements and identify prominent research questions that remain, a workshop was organized by IARC and the US NCI. A series of 'difficult questions' were formulated, which should be given future priority in the areas of population, genomic and clinical research.

Mechanical signatures of microbial biofilms in micropillar-embedded growth chambers.

Biofilms are surface-attached communities of microorganisms embedded in an extracellular matrix and are essential for the cycling of organic matter in natural and engineered environments. They are also the leading cause of many infections, for example, those associated with chronic wounds and implanted medical devices. The extracellular matrix is a key biofilm component that determines its architecture and defines its physical properties. Herein, we used growth chambers embedded with micropillars to study the net mechanical forces (differential pressure) exerted during biofilm formation in situ. Pressure from the biofilm is transferred to the micropillars via the extracellular matrix, and reduction of major matrix components decreases the magnitude of micropillar deflections. The spatial arrangement of micropillar deflections caused by pressure differences in the different biofilm strains may potentially be used as mechanical signatures for biofilm characterization. Hence, we submit that micropillar-embedded growth chambers provide insights into the mechanical properties and dynamics of the biofilm and its matrix.

A proposal for a new scoring system to predict difficult ventilation through a supraglottic airway.

BACKGROUND: The aim of this study was to propose and validate a new clinical score to predict difficult ventilation through a supraglottic airway device. METHODS: The score was proposed from our previously reported derivation data, and we prospectively validated the score in 5532 patients from November 2013 to April 2014. Predictive accuracy of the score was compared by the area under the receiver operating characteristic (ROC) curve (AUC). We assigned point values to each of the identified four risk factors: male, age >45 yr, short thyromental distance, and limited neck movement, their sum composing the score. The score ranged between 0 and 7 points. The optimal predictive level of the score was determined using ROC curve analysis. RESULTS: The AUC of the score was 0.75 (95% CI 0.66 to 0.84) in the validation data set, and was similar to that in the derivation data set (0.80; 95% CI 0.75 to 0.86). In derivation and validation data sets, the incidence of low risk categories (scores 0-3) was 0.42% vs 0.32% and of high risk categories (scores 4-7) was 3% vs 1.7% respectively. A score 4 or greater is associated with a six to seven fold increased risk of difficult ventilation through a supraglottic airway device. CONCLUSIONS: The new score for prediction of difficult ventilation through a supraglottic airway device is easy to perform and reliable, and could help anaesthetists plan for difficult airway management.

Miniaturized versus conventional cardiopulmonary bypass and acute kidney injury after cardiac surgery.

INTRODUCTION: Acute kidney injury (AKI) is a serious complication after coronary artery bypass grafting (CABG). There are conflicting reports whether a miniaturized cardiopulmonary bypass (MCPB) system is associated with a lower AKI incidence compared with conventional cardiopulmonary bypass (CCPB). It is unknown if AKI risk factors differ between the two groups. We assessed if MCPB decreases AKI after CABG and compared the risk factors between both groups. METHODS: Sixty-eight Asian patients presenting for elective CABG at a tertiary heart centre were enrolled. They were randomly assigned to MCPB (n=34) or CCPB group (n=34) and followed up in a single-blinded, prospective, randomized, controlled trial. The primary outcome was Acute Kidney Injury Network stage 1 AKI. RESULTS: The AKI incidence was 21.5% and was not significantly different between patients undergoing MCPB versus CCPB (21.9% versus 21.2%, p=0.948). The first CPB haematocrit was independently associated with AKI in the MCPB group (Relative Risk [RR]=0.484, 95% Confidence Interval [CI]=0.268-0.876, p=0.016); post-operative blood loss and inflammation were independently associated with AKI in the CCPB group (RR=1.005, 95%CI=1.003-1.007, p<0.001; RR=1.018, 95%CI=1.010-1.028, p<0.001). CONCLUSION: The MCPB system is not associated with a lower incidence of AKI in Asian patients undergoing CABG. Risk factors for AKI differed between patients using the MCPB and CCPB systems.

Geographic differences in the associations between impaired glucose regulation and cardiovascular risk factors among young adults.

AIMS: To assess geographic differences in the association between BMI, blood pressure and lipid levels with impaired glucose regulation among young adults from various geographical regions. METHODS: This was a cross-sectional study including data from 6987 participants aged ≤ 30 years from India, Singapore, Australia, Greenland, Kenya and Tanzania. Impaired glucose regulation was determined by the 75-g oral glucose tolerance test. For each geographical region, BMI, blood pressure and lipids were examined and compared between participants with normal glucose tolerance and those with impaired glucose regulation. Multiple logistic regression models were used to assess the association between risk factors and impaired glucose regulation. RESULTS: Indian and East African people had a higher prevalence of impaired glucose regulation compared with participants from other regions, despite their lower BMI. Compared with the other regions, blood pressure was lower among Indian and Singaporean people but higher in those from Greenland. Greenlanders had the highest, while Indian and East-African people, had the lowest level of HDL cholesterol. BMI was positively associated with impaired glucose regulation in all regions, and there were no statistically significant geographic differences. In the Indian, Singaporean and Australian participants, there was a positive association between blood pressure and impaired glucose regulation. Triglycerides were positively associated with and HDL cholesterol had no association with impaired glucose regulation in all geographical regions. CONCLUSIONS: Higher BMI and triglyceride levels were positively associated with prevalent impaired glucose regulation in all geographical regions. There were geographic differences in the association between impaired glucose regulation and blood pressure and lipids, probably reflecting environmental and genetic factors.

Signal molecule changes in the gills and lungs of the African lungfish Protopterus annectens, during the maintenance and arousal phases of aestivation.

African lungfishes are obligate air breathers, with reduced gills and pulmonary breathing throughout their life. During the dry season they aestivate on land, with the collapse of secondary lamellae of their gills and the establishment of an exclusive aerial ventilation through the vascularization and expansion of their lungs. To date, the mechanisms underlining the respiratory organ remodeling in aestivating lungfishes are unknown. This study aimed to identify key switch components of the stress-induced signal transduction networks implicated in both rapid and medium-long term remodeling of the gills and lungs of the African lungfish Protopterus annectens during aestivation. Through immunofluorescence microscopy and Western blotting, the localization and the expression of nitric oxide synthase (NOS), Akt, Hsp-90 and HIF-1α were evaluated in both gills and lungs exposed to three experimental conditions: freshwater (FW), 6 months of experimentally induced aestivation (6mAe), and 6 days after arousal from 6 months of aestivation (6mAe6d). After 6mAe, the expression of NOS (p-eNOS antibody), Akt (p-Akt antibody), and Hsp-90 decreased in the gills, while NOS and Hsp-90 expression increased with Akt remained unchanged in the lungs. Upon 6mAe6d, NOS, Akt and Hsp-90 expression in the gills returned to the respective FW values. In the lungs of the aroused fish, NOS and Akt decreased to their respective FW levels, while Hsp-90 expression was enhanced with respect to aestivation. In both respiratory organs, the qualitative and quantitative patterns of HIF-1α expression correlated inversely to those of NOS. Overall, our findings suggest that the molecular components of the NOS/NO system changed in a tissue-specific manner in parallel with organ readjustment in the gills and lungs of P. annectens during aestivation and arousal.

Probiotic Lactobacillus rhamnosus GR-1 and Lactobacillus reuteri RC-14 exhibit strong antifungal effects against vulvovaginal candidiasis-causing Candida glabrata isolates.

AIMS: This study investigates the antagonistic effects of the probiotic strains Lactobacillus rhamnosus GR-1 and Lactobacillus reuteri RC-14 against vulvovaginal candidiasis (VVC)-causing Candida glabrata. METHODS AND RESULTS: Growth inhibitory activities of Lact. rhamnosus GR-1 and Lact. reuteri RC-14 strains against C. glabrata were demonstrated using a spot overlay assay and a plate-based microtitre assay. In addition, these probiotic lactobacilli strains also exhibited potent candidacidal activity against C. glabrata, as demonstrated by a LIVE/DEAD yeast viability assay performed using confocal laser scanning microscopy. The metabolic activities of all C. glabrata strains were completely shut down in response to the challenges by the probiotic lactobacilli strains. In addition, both probiotic lactobacilli strains exhibited strong autoaggregation and coaggregation phenotypes in the presence of C. glabrata, which indicate that these lactobacilli strains may exert their probiotic effects through the formation of aggregates and, thus the consequent prevention of colonization by C. glabrata. CONCLUSIONS: Probiotic Lact. rhamnosus GR-1 and Lact. reuteri RC-14 strains exhibited potent antagonistic activities against all of the tested C. glabrata strains. These lactobacilli exhibited antifungal effects, including those attributed to their aggregation abilities, and their presence caused the cessation of growth and eventual cell death of C. glabrata. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study to report on the antagonistic effects of these probiotic lactobacilli strains against the non-Candida albicans Candida (NCAC) species C. glabrata.

Incidence of and risk factors for difficult ventilation via a supraglottic airway device in a population of 14,480 patients from South-East Asia.

Difficult airway practice guidelines include the use of a supraglottic airway device as part of the armamentarium to provide and maintain ventilation and oxygenation. We retrospectively reviewed 14 480 patients aged ≥ 18 years who underwent general anaesthesia. We identified 74 (0.5%) patients whose lungs were identified as having been difficult to ventilate via a supraglottic airway device, and 29 (0.2%) patients in whom device placement failed. Multivariate analysis identified four risk factors for difficult ventilation via a supraglottic airway device: male sex (OR 1.75, 95% CI 1.07-2.86, p = 0.02); age > 45 years (OR 1.70, 95% CI 1.01-2.86, p = 0.04); short thyromental distance (OR 4.35, 95% CI 2.31-8.17, p < 0.001); and limited neck movement (OR 2.75, 95% CI 1.02-7.44, p = 0.04). Adverse respiratory events including oxygen desaturation, hypercapnoea, laryngospasm, and bronchospasm occurred in 17 patients (22%). The incidence of difficult ventilation via a supraglottic airway device was 0.5% in a large cohort of South-East Asian patients.

The inflammatory response between miniaturised and conventional cardiopulmonary bypass after cardiac surgery in an Asian population.

INTRODUCTION: We compared the systemic inflammatory response of the MCPB system to the CCPB system with cell salvage and phosphorylcholine-coated tubing amongst Asian patients undergoing coronary artery bypass grafting. METHODS: Seventy-eight patients were randomly assigned to the MCPB or the CCPB groups equally and followed up in a prospective, single-blinded, randomised, controlled trial. Levels of TNF-α, IL-6, CRP and LDH were measured peri-operatively. RESULTS: The systemic inflammatory response was similar in both groups (TNF-α: p=0.222; IL-6: p=0.991; CRP: p=0.258). Only haemolysis was significantly higher in the CCPB group (LDH: p=0.011). The MCPB system was twice more expensive, but had a near 4-fold cost saving in tranfusions. Overall, the MCPB system cost 20% more than the modified CCPB system. CONCLUSION: These results corroborate with studies that demonstrated the avoidance of cardiotomy suction rather than the MCPB system, itself, leads to an attenuated inflammatory response. The absence of obvious clinical benefit and the higher costs involved with the MCPB system would preclude its routine use.

Excretory nitrogen metabolism and defence against ammonia toxicity in air-breathing fishes.

With the development of air-breathing capabilities, some fishes can emerge from water, make excursions onto land or even burrow into mud during droughts. Air-breathing fishes have modified gill morphology and morphometry and accessory breathing organs, which would tend to reduce branchial ammonia excretion. As ammonia is toxic, air-breathing fishes, especially amphibious ones, are equipped with various strategies to ameliorate ammonia toxicity during emersion or ammonia exposure. These strategies can be categorized into (1) enhancement of ammonia excretion and reduction of ammonia entry, (2) conversion of ammonia to a less toxic product for accumulation and subsequent excretion, (3) reduction of ammonia production and avoidance of ammonia accumulation and (4) tolerance of ammonia at cellular and tissue levels. Active ammonia excretion, operating in conjunction with lowering of ambient pH and reduction in branchial and cutaneous NH3 permeability, is theoretically the most effective strategy to maintain low internal ammonia concentrations. NH3 volatilization involves the alkalization of certain epithelial surfaces and requires mechanisms to prevent NH3 back flux. Urea synthesis is an energy-intensive process and hence uncommon among air-breathing teleosts. Aestivating African lungfishes detoxify ammonia to urea and the accumulated urea is excreted following arousal. Reduction in ammonia production is achieved in some air-breathing fishes through suppression of amino acid catabolism and proteolysis, or through partial amino acid catabolism leading to alanine formation. Others can slow down ammonia accumulation through increased glutamine synthesis in the liver and muscle. Yet, some others develop high tolerance of ammonia at cellular and tissue levels, including tissues in the brain. In summary, the responses of air-breathing fishes to ameliorate ammonia toxicity are many and varied, determined by the behaviour of the species and the nature of the environment in which it lives.

Nitric oxide synthase-dependent "on/off" switch and apoptosis in freshwater and aestivating lungfish, Protopterus annectens: skeletal muscle versus cardiac muscle.

African lungfishes (Protopterus spp.) are obligate air breathers which enter in a prolonged torpor (aestivation) in association with metabolic depression, and biochemical and morpho-functional readjustments during the dry season. During aestivation, the lungfish heart continues to pump, while the skeletal muscle stops to function but can immediately contract during arousal. Currently, nothing is known regarding the orchestration of the multilevel rearrangements occurring in myotomal and myocardial muscles during aestivation and arousal. Because of its universal role in cardio-circulatory and muscle homeostasis, nitric oxide (NO) could be involved in coordinating these stress-induced adaptations. Western blotting and immunofluorescence microscopy on cardiac and skeletal muscles of Protopterus annectens (freshwater, 6months of aestivation and 6days after arousal) showed that expression, localization and activity of the endothelial-like nitric oxide synthase (eNOS) isoform and its partners Akt and Hsp-90 are tissue-specifically modulated. During aestivation, phospho-eNOS/eNOS and phospho-Akt/Akt ratios increased in the heart but decreased in the skeletal muscle. By contrast, Hsp-90 increased in both muscle types during aestivation. TUNEL assay revealed that increased apoptosis occurred in the skeletal muscle of aestivating lungfish, but the myocardial apoptotic rate of the aestivating lungfish remained unchanged as compared with the freshwater control. Consistent with the preserved cardiac activity during aestivation, the expression of apoptosis repressor (ARC) also remained unchanged in the heart of aestivating and aroused fish as compared with the freshwater control. Contrarily, ARC expression was strongly reduced in the skeletal muscle of aestivating lungfish. On the whole, our data indicate that changes in the eNOS/NO system and cell turnover are implicated in the morpho-functional readjustments occurring in lungfish cardiac and skeletal muscle during the switch from freshwater to aestivation, and between the maintenance and arousal phases of aestivation.