Rapid and specific determination of DNA adducts for clinical diagnosis of poisoning and disease associated with aristolochic acid.

1. A liquid chromatography-mass spectrometry method was developed to detect and characterise aristocholic acid-DNA adducts in biological samples. 2. The detection of DNA adducts in plasma, urine or the cells found in urine may be useful to support the diagnosis and monitoring of aristocholic acid-associated poisoning and disease. 3. Efforts should be made to improve the sensitivity and specificity of this approach for the detection and characterisation of exposure to other mutagens/carcinogens.

Prevalence of vertebral fracture in Asian men and women: comparison between Hong Kong, Thailand, Indonesia and Japan.

OBJECTIVES: Little is known about the prevalence of vertebral fracture among Asians. This study investigated the prevalence of radiographically defined vertebral fracture, and identified associated risk factors in the aged population of four Asian countries. STUDY DESIGN: In total, 1588 males and females aged ≥ 65 years were recruited from Hong Kong, Thailand, Indonesia and Japan. METHODS: Standard X-rays for the spine were taken and vertebral heights were measured. Vertebral fracture was defined as a reduction of >3 standard deviations in vertebral height ratio. Bone mineral density (BMD) of the hip was measured by dual energy X-ray absorptiometry, and anthropometric measurements were taken in Hong Kong and Japan. Other relevant data were entered in a standard questionnaire. RESULTS: The prevalence of vertebral fracture for both males and females was highest in Japan for younger (65-74 years) and older (≥ 75 years) age groups (36.6% and 37.6% for males; 18.8% and 28.7% for females). Lower hip BMD was associated with vertebral fracture in both sexes. Older age, lower quality of life score on Short Form-12 (physical), past longest occupation as a farmer, and history of cataract were significantly associated with vertebral fracture in females. However, smoking did not appear to be an important risk factor for vertebral fracture. CONCLUSIONS: Radiographic assessments for vertebral fracture were performed in all four Asian countries. The prevalence of vertebral fracture was highest in Japan. Lower hip BMD, poorer physical condition and past longest occupation as a farmer were associated with vertebral fracture.

Using the Theory of Planned Behaviour and implementation intentions to predict and facilitate upward family communication about mammography.

Regular mammography facilitates early detection of breast cancer, and thus increases the chances of survival from this disease. Daughter-initiated (i.e. upward) communication about mammography within mother-daughter dyads may promote mammography to women of screening age. The current study examined this communication behaviour within the context of the Theory of Planned Behaviour (TPB), and aimed to bridge the intention-behaviour gap by trialling an implementation intention (II) intervention that aimed to facilitate upward family communication about mammography. Young women aged 18-39 (N=116) were assigned to either a control or experimental condition, and the latter group formed IIs about initiating a conversation with an older female family member about mammography. Overall, those who formed IIs were more likely to engage in the target communication behaviour, however the intervention was most effective for those who reported low levels of intention at baseline. Perceived behavioural control emerged as the most important variable in predicting the target behaviour. The altruistic nature of this behaviour, and the fact that it is not wholly under volitional control, may have contributed to this finding. Future studies that systematically explore the relative roles of intention and perceived behavioural control in behaviours of this nature are warranted.

BAG3-related myofibrillar myopathy in a Chinese family.

In contrast to the usual slow disease progression in myofibrillar myopathies, patients with Bag3opathy often have a rapidly progressive and more severe phenotype with a worse prognosis. We describe a Chinese patient, born to non-consanguineous parents, who first presented at age 6 with clumsy walking and difficult climbing staircase. With a history of restrictive lung disease previously diagnosed as asthma, she progressed rapidly with proximal myopathy, rigid spine and bilateral tightening of the Achilles tendons requiring surgical elongation. Hypertrophic cardiomyopathy with restrictive physiology was shown by echocardiogram. Moreover, prolonged QT interval was also noted in the patient. Family history was unremarkable yet her father was incidentally found to have prolonged QT interval. Mutation analysis with genomic DNA of the proband showed heterozygous de novo known mutation c.626C>T (p.Pro209Leu) and a germline variation c.772C>T (p.Arg258Trp) in BAG3. Her father was found to be a carrier of c.772C>T. Muscle biopsy findings were suggestive of myofibrillar myopathy on light microscopy and ultrastructural studies. To our knowledge, this is the first Chinese case of Bag3opathy so far reported.

RYR1-related central core myopathy in a Chinese adolescent boy.

Central core myopathy is a rare, inherited neuromuscular disorder with a wide spectrum of phenotypic presentations. It is also considered an allelic disease of malignant hyperthermia. We report a case of central core myopathy in a Chinese adolescent boy presenting with atypical clinical features and a moderately elevated serum creatine kinase level. The diagnosis was made from the histopathological findings of central cores on muscle biopsy, and confirmed by the molecular genetic testing for the RYR1 gene mutation. This is the first case of central core myopathy confirmed by molecular study in our locality.

Refractory bulbar and respiratory dysfunction in a young Chinese woman with seronegative, muscle-specific tyrosine kinase antibody-positive myasthenia gravis: response to cyclophosphamide and rituximab treatment.

The use of cyclophosphamide and rituximab for patients with refractory myasthenia gravis has shown promising results. We report on a 31-year-old Chinese woman with acetylcholine receptor antibody-negative and muscle-specific tyrosine kinase antibody-positive generalised myasthenia gravis who had refractory bulbar dysfunction and respiratory failure despite immunosuppressive therapy and thymectomy, and partial and sustained responses to cyclophosphamide and rituximab treatment, respectively. Myasthenia crisis was diagnosed when she presented in the third trimester of pregnancy with dysphagia, bilateral ptosis, prominent fatigability, and respiratory failure. She required prolonged intensive care and non-invasive ventilatory support despite several courses of intravenous immunoglobulins and plasmapheresis. Pulse cyclophosphamide 500 mg/m(2) was given monthly for 4 consecutive months with a partial response. Rituximab 500 mg weekly was subsequently given for 4 weeks with a dramatic and sustained response. She remained symptom-free and assumed full maternal care at 1 year. To the authors' knowledge, this is the first report of a Chinese patient with refractory myasthenia gravis who responded to cyclophosphamide and rituximab.

Macroprolactin-a cause of pseudohyperprolactinaemia.

Macroprolactin is a complex of immunoglobulin G and monomeric prolactin with little biological activity in vivo. Macroprolactin cross-reacts in modern commercial prolactin assays, however, leading to pseudohyperprolactinaemia. This report is of three patients with macroprolactinaemia and the untoward consequences if this benign condition is misdiagnosed as genuine hyperprolactinaemia are discussed. One adult and one child without symptoms of hyperprolactinaemia were incidentally found to have elevated serum prolactin levels, one of whom had a pituitary incidentaloma. Repeat prolactin measurement after polyethylene glycol precipitation showed that the majority of circulating prolactin was macroprolactin. The third patient had galactorrhoea and pituitary microadenoma. Polyethylene glycol study showed that macroprolactinaemia exists simultaneously with genuine hyperprolactinaemia leading to falsely high serum prolactin levels. The recognition of this relatively common and benign condition is important in order to avoid misdiagnosis and unnecessary investigations and treatment. Particular attention must be paid to patients in whom the clinical and radiological findings are incompatible.

Infantile isolated sulphite oxidase deficiency in a Chinese family: a rare neurodegenerative disorder.

We report the clinical, biochemical, neuroradiological, and neurophysiological findings of a 4-year-old Chinese girl with infantile isolated sulphite oxidase deficiency. This is the first reported case in our locality. She presented at the age of 5 months with refractory seizures and developmental regression, and progressed rapidly to profound psychomotor retardation, spasticity, dystonia, microcephaly, and blindness. At the age of 3.5 years, she was admitted to the intensive care unit with septic shock. Ophthalmologic examination at this time revealed bilateral dislocation of the lens. Diagnosis of this very rare disorder was made on the basis of increased levels of urinary sulphite, thiosulphate, and sulphocysteine; normal urine xanthine and hypoxanthine; normal plasma uric acid; and low plasma cystine levels. The diagnosis was confirmed by the absence of sulphite oxidase activities in skin fibroblasts. Isolated sulphite oxidase deficiency is a rare inborn error of sulphur metabolism that is difficult to diagnose on clinical features and routine metabolic tests. The presence of ectopia lentis, seizures, and progressive neurological abnormalities should alert clinicians to the diagnosis.

Platelet Na,K-adenosine triphosphatase as a tissue marker of hyperthyroidism.

Platelet Na(+),K(+)-adenosine triphosphatase (ATPase) activity was measured in 34 (15 males, 19 females) healthy subjects, 89 (35 males, 54 females) hyperthyroid patients, and 34 (7 males, 27 females) treated hyperthyroid patients to assess the potential of this measurement as a tissue marker and diagnostic test for hyperthyroidism. Platelet Na(+),K(+)-ATPase activity was measured in platelet lysates by the rate of release of phosphate from adenosine triphosphate (ATP) in the presence and absence of ouabain. Platelet Na(+),K(+)-ATPase activity (median and range) in the hyperthyroid group (271, 169 to 821 pmol/h/g protein) was significantly higher compared with the healthy group (125, 74 to 185 micromol/h/g protein, P <.001 by Mann-Whitney U test). The treated hyperthyroid group had slightly, but significantly higher, free triiodothyronine (FT3) and free thyroxine (FT4), as well as platelet Na(+),K(+)-ATPase activity (147, 98 to 246 micromol/h/g protein, P <.05). If a platelet Na(+),K(+)-ATPase activity of 190 micromol/h/g protein was used as a cut off value, the specificity and sensitivity were 90% and 93%, respectively. We conclude that platelet Na(+),K(+)-ATPase may be a useful tissue marker of hyperthyroidism.

Should we screen for globin gene mutations in blood samples with mean corpuscular volume (MCV) greater than 80 fL in areas with a high prevalence of thalassaemia?

AIMS: To investigate whether it is worthwhile, in areas where thalassaemia is common, to screen for globin gene mutations in subjects with a mean corpuscular volume (MCV) above 80 fL, especially in partners of known thalassaemia carriers. METHODS: Blood samples from 95 subjects with MCV between 80 and 85 fL were screened for the presence of alpha globin gene mutations and the haemoglobin (Hb) E mutation. RESULTS: Thirty four subjects harboured globin gene mutations. Of these, 31 had deletions of one alpha globin gene, one had Hb Constant Spring, and three had Hb E mutations. CONCLUSION: Based on the above figures and known prevalence rates of thalassaemia carriers, it would seem worthwhile to screen for globin gene mutations in partners of known thalassaemia carriers, regardless of MCV, to identify pregnancies at risk of Hb H disease or Hb E/beta thalassaemia.

Outcome of 85 lupus nephritis patients treated with intravenous cyclophosphamide: a single centre 10 year experience.

Retrospective analysis was done on 85 patients (76 female, 9 male) with lupus nephritis who started intravenous cyclophosphamide between 1/1/1989 and 31/12/1998. The initial renal biopsy (World Health Organisation) classification was III (4.7%), IV (89.4%) and V (5.9%). Average serum creatinine at time of biopsy was 0.12 +/- 0.12 mmol/l. Median duration of nephritis before biopsy was 2 months (range 0-133). Median duration of follow-up from time of biopsy to outcome (death or end-stage renal failure) was 3.3 years (range 0.3-11.8). Nineteen patients died. The calculated proportion alive at 5 years was 75% and at 10 years 64%. The calculated proportion alive with renal function was 74% and 54% at 5 and 10 years respectively. Fifty-two patients completed cyclophosphamide therapy at the end of the study. There were ten episodes of herpes zoster, the most common infection seen. No malignancy was reported.

Recurrent upper gastrointestinal bleeding due to the Dieulafoy's lesion.

Dieulafoy's lesion of the gastrointestinal tract, an abnormally dilated artery that penetrates through the mucosa, has been diagnosed more frequently in recent years. Bleeding occurs when the vessel ruptures. Despite widespread awareness of this entity, it remains a diagnostic challenge for gastroenterologists because of its small size and hidden location. Several different diagnostic methods and treatments have been introduced throughout the years. Endoscopy plays a major role in diagnosis and therapy. In this case report, the patient was successfully treated endoscopically using epinephrine injection and heater probe thermoco-agulation. The characteristics of the Dieulafoy's lesion, its current diagnosis, treatment, and complications are discussed.

Role of cofilin in epidermal growth factor-stimulated actin polymerization and lamellipod protrusion.

Stimulation of metastatic MTLn3 cells with epidermal growth factor (EGF) causes a rapid and transient increase in actin nucleation activity resulting from the appearance of free barbed ends at the extreme leading edge of extending lamellipods. To investigate the role of cofilin in EGF-stimulated actin polymerization and lamellipod extension in MTLn3 cells, we examined in detail the temporal and spatial distribution of cofilin relative to free barbed ends and characterized the actin dynamics by measuring the changes in the number of actin filaments. EGF stimulation triggers a transient increase in cofilin in the leading edge near the membrane, which is precisely cotemporal with the appearance of free barbed ends there. A deoxyribonuclease I binding assay shows that the number of filaments per cell increases by 1.5-fold after EGF stimulation. Detection of pointed ends in situ using deoxyribonuclease I binding demonstrates that this increase in the number of pointed ends is confined to the leading edge compartment, and does not occur within stress fibers or in the general cytoplasm. Using a light microscope severing assay, cofilin's severing activity was observed directly in cell extracts and shown to be activated after stimulation of the cells with EGF. Microinjection of function-blocking antibodies against cofilin inhibits the appearance of free barbed ends at the leading edge and lamellipod protrusion after EGF stimulation. These results support a model in which EGF stimulation recruits cofilin to the leading edge where its severing activity is activated, leading to the generation of short actin filaments with free barbed ends that participate in the nucleation of actin polymerization.

Tetrahydropalmatine poisoning: diagnoses of nine adult overdoses based on toxicology screens by HPLC with diode-array detection and gas chromatography-mass spectrometry.

BACKGROUND: Tetrahydropalmatine (THP) is a neuroactive alkaloid with analgesic and hypnotic action. Its analysis is important because cases of human poisonings have emerged as a result of unregulated use of some proprietary biopharmaceuticals containing purified THP. METHODS: We established analytical parameters for HPLC with diode-array detection (HPLC-DAD) and gas chromatography-mass spectrometry (GC-MS) for the detection of THP in serum and urine. Nine acutely THP-poisoned adults were thus screened over 16 months. RESULTS: All patients recovered quickly after mild neurological disturbance. In general, THP was metabolized rapidly and excreted as polar metabolites in urine. Serum THP was measured in five cases and found to be <0.1-1.2 mg/L (<0.3-3.4 micromol/L). Paired analyses of urine with and without glucuronidase treatment clarified the disposition of THP. Our GC-MS method with trimethylsilane derivatization identified O-desmethyl metabolites. With a uniform solid-phase extraction, the HPLC-DAD procedure detected intact glucuronide metabolites. CONCLUSION: Intact glucuronide metabolites of THP are sensitive markers for THP exposures. Our methods and findings provide practical tools and information for surveillance of intoxication caused by excessive THP intake.

Myelodysplastic syndrome with myelofibrosis and basophilia: detection of trisomy 8 in basophils by fluorescence in-situ hybridization.

We report a case of myelodysplastic syndrome (MDS) with myelofibrosis and pulmonary tuberculosis who had marked basophilia in the peripheral blood. A clonal karyotypic abnormality characterized by trisomy 8 was demonstrated by cytogenetic analysis. By correlation of cell morphology with results of fluorescence in situ hybridization using a chromosome 8 probe, we demonstrated that the basophils were not reactive but belonged to the neoplastic MDS clone.

Intraperitoneal pulse calcitriol in the treatment of secondary hyperparathyroidism in patients on continuous ambulatory peritoneal dialysis.

OBJECTIVE: To determine whether it is practicable to use intraperitoneal calcitriol to treat continuous ambulatory peritoneal dialysis (CAPD) patients with secondary hyperparathyroidism and whether this form of therapy is effective and safe. DESIGN: A prospective, nonrandomized study. SETTING: Division of Nephrology, Tertiary Hospital. METHOD: Eight patients from our CAPD population were recruited (5 F, 3 M), aged 24 to 63 years (mean 38.9 +/- 7.6 yr). They had been on CAPD for 8 to 84 months (mean 47.6 +/- 24.6 months). All the patients had bone biopsy-proven secondary hyperparathyroidism with 2 patients showing mixed lesions. The CAPD system was changed to the twin-bag system (Ultrabag, Baxter Healthcare, McGaw Park, IL, U.S.A.) in all 8 patients, who were taught to inject the calcitriol directly through the short transfer set and the Tenckhoff catheter into the peritoneal cavity, twice per week before bedtime. Calcium carbonate or calcium acetate was used as the main phosphate binder. Intact parathyroid hormone level (iPTH), serum ionized calcium (iCa), serum phosphate, and serum total alkaline phosphatase (alk. phos.) levels were measured at baseline and then every 4 weeks. The mean duration of follow-up was 10.5 months +/- 1.9 months. RESULTS: There was a significant drop of iPTH level from the pretreatment level of 100.6 +/- 35.8 pmol/L to a level of 63.8 +/- 48.7 pmol/L at 24 weeks (p = 0.036). The lowest level of iPTH attained was 43.4 +/- 27.0 pmol/L at 48 weeks. Serum total alk. phos. also dropped from 232.4 +/- 83.3 IU/L pretreatment to 147.9 +/- 52.0 IU/L at 24 weeks (p = 0.017). The decrease in alk. phos. level paralleled the decrease in iPTH level. The mean serum iCa level did not show any significant rise throughout the study period. The maximum dose of calcitriol used was 6.6 +/- 1.5 microg/week and the average dose of calcitriol was 5.4 +/- 1.2 microg/week. One patient did not respond satisfactorily and she subsequently had a parathyroidectomy. Two episodes of peritonitis occurred during the study period, giving a peritonitis rate of one episode per 42 patient-months. There was no significant change in the urea clearance tests or the peritoneal equilibration tests before and after the study. CONCLUSION: Intraperitoneal calcitriol is practicable, effective, and safe in the treatment of secondary hyperparathyroidism in CAPD patients.

EGF stimulates an increase in actin nucleation and filament number at the leading edge of the lamellipod in mammary adenocarcinoma cells.

Stimulation of metastatic MTLn3 cells with EGF causes the rapid extension of lamellipods, which contain a zone of F-actin at the leading edge. In order to establish the mechanism for accumulation of F-actin at the leading edge and its relationship to lamellipod extension in response to EGF, we have studied the kinetics and location of EGF-induced actin nucleation activity in MTLn3 cells and characterized the actin dynamics at the leading edge by measuring the changes at the pointed and barbed ends of actin filaments upon EGF stimulation of MTLn3 cells. The major result of this study is that stimulation of MTLn3 cells with EGF causes a transient increase in actin nucleation activity resulting from the appearance of free barbed ends very close to the leading edge of extending lamellipods. In addition, cytochalasin D causes a significant decrease in the total F-actin content in EGF-stimulated cells, indicating that both actin polymerization and depolymerization are stimulated by EGF. Pointed end incorporation of rhodamine-labeled actin by the EGF stimulated cells is 2.12+/-0.47 times higher than that of control cells. Since EGF stimulation causes an increase in both barbed and pointed end incorporation of rhodamine-labeled actin in the same location, the EGF-stimulated nucleation sites are more likely due either to severing of pre-existing filaments or de novo nucleation of filaments at the leading edge thereby creating new barbed and pointed ends. The timing and location of EGF-induced actin nucleation activity in MTLn3 cells can account for the observed accumulation of F-actin at the leading edge and demonstrate that this F-actin rich zone is the primary actin polymerization zone after stimulation.