Detection of SYT-SSX mutant transcripts in formalin-fixed paraffin-embedded sarcoma tissues using one-step reverse transcriptase real-time PCR.

BACKGROUND: Synovial sarcoma (SS) is a rare cancer and accounts for 5-10% of adult soft tissue sarcomas. Making an accurate diagnosis is difficult due to the overlapping histological features of SS with other types of sarcomas and the non-specific immunohistochemistry profile findings. Molecular testing is thus considered necessary to confirm the diagnosis since more than 90% of SS cases carry the transcript of t(X;18)(p11.2;q11.2). The purpose of this study is to diagnose SS at molecular level by testing for t(X;18) fusion-transcript expression through One-step reverse transcriptase real-time Polymerase Chain Reaction (PCR). METHOD: Formalin-fixed paraffin-embedded tissue blocks of 23 cases of soft tissue sarcomas, which included 5 and 8 cases reported as SS as the primary diagnosis and differential diagnosis respectively, were retrieved from the Department of Pathology, Tengku Ampuan Afzan Hospital, Kuantan, Pahang. RNA was purified from the tissue block sections and then subjected to One-step reverse transcriptase real-time PCR using sequence specific hydrolysis probes for simultaneous detection of either SYT-SSX1 or SYT-SSX2 fusion transcript. RESULTS: Of the 23 cases, 4 cases were found to be positive for SYT-SSX fusion transcript in which 2 were diagnosed as SS whereas in the 2 other cases, SS was the differential diagnosis. Three cases were excluded due to failure of both amplification assays SYT-SSX and control ß-2-microglobulin. The remaining 16 cases were negative for the fusion transcript. CONCLUSION: This study has shown that the application of One-Step reverse transcriptase real time PCR for the detection SYT-SSX transcript is feasible as an aid in confirming the diagnosis of synovial sarcoma.

Detection of SYT-SSX mutant transcripts in formalin-fixed paraffin-embedded sarcoma tissues using one-step reverse transcriptase real-time PCR

Background: Synovial sarcoma (SS) is a rare cancer and accounts for 5-10% of adult soft tissue sarcomas. Making an accurate diagnosis is difficult due to the overlapping histological features of SS with other types of sarcomas and the non-specific immunohistochemistry profile findings. Molecular testing is thus considered necessary to confirm the diagnosis since more than 90% of SS cases carry the transcript of t(X;18)(p11.2;q11.2). The purpose of this study is to diagnose SS at molecular level by testing for t(X;18) fusion-transcript expression through One-step reverse transcriptase real-time Polymerase Chain Reaction (PCR). Method: Formalin-fixed paraffin-embedded tissue blocks of 23 cases of soft tissue sarcomas, which included 5 and 8 cases reported as SS as the primary diagnosis and differential diagnosis respectively, were retrieved from the Department of Pathology, Tengku Ampuan Afzan Hospital, Kuantan, Pahang. RNA was purified from the tissue block sections and then subjected to One-step reverse transcriptase real-time PCR using sequence specific hydrolysis probes for simultaneous detection of either SYT-SSX1 or SYT-SSX2 fusion transcript. Results: Of the 23 cases, 4 cases were found to be positive for SYT-SSX fusion transcript in which 2 were diagnosed as SS whereas in the 2 other cases, SS was the differential diagnosis. Three cases were excluded due to failure of both amplification assays SYT-SSX and control β-2-microglobulin. The remaining 16 cases were negative for the fusion transcript. Conclusion: This study has shown that the application of One-Step reverse transcriptase real time PCR for the detection SYT-SSX transcript is feasible as an aid in confirming the diagnosis of synovial sarcoma.

Inherited Anti-Thrombin Deficiency in A Malay-Malaysian Family: A Missense Mutation at Nucleotide g.13267C>A aka anti-thrombin Budapest 5 (p.Pro439Thr) of the SERPINC 1 gene.

OBJECTIVE: Inherited anti-thrombin deficiency is an autosomal dominant disorder which is associated with increased risk for venous thromboembolism (VTE). This condition is very rare in Malaysia and there has been no documented report. Thus, the aim of the present study is to investigate the type of an inherited anti-thrombin deficiency mutation in a 25-year-old Malay woman who presented with deep vein thrombosis in her first pregnancy. METHODS: DNA was extracted from the patient's blood sample and buccal mucosal swabs from family members. Polymerase chain reaction(PCR) assays were designed to cover all seven exons of the serpin peptidase inhibitor, clade C (antithrombin), member 1 (SERPINC1) gene; and the products were subjected to DNA sequencing. Sequences were referred to NCBI Reference Sequence: NG_012462.1. RESULTS: A heterozygous substitution mutation at nucleotide position 13267 (CCT->ACT) was identified in the patient and two other family members, giving a possible change of codon 439 (Pro→Thr) also known as anti-thrombin Budapest 5. The genotype was absent in 90 healthy controls. CONCLUSION: The study revealed a heterozygous antithrombin Budapest 5 mutation in SERPINC 1 giving rise to a possible anti-thrombin deficiency in a Malay-Malaysian family.

Study the live weight and live weight gain of black bengal and jamunapari goat breeds by fitting the linear regression under semi-intensive conditions.

The present study was conducted to know the live weight gain of goats under semi-intensive conditions of Chittagong district of Bangladesh during the period of July, 2012 to January, 2013. Data were collected from 72 black Bengal and 32 Jamunapari goats and the kids birth weight and their subsequent live weight at weekly intervals up to age and weight of weaning at sexual maturity was recorded. The weight gains from birth to sexual maturity of two different breeds under 2 different farms were studied. Average birth weight of male and female black Bengal goats kids were 1.22 +/- 0.15, 1.01 +/- 0.14, 1.42 +/- 0.10 and 1.12 +/- 0.27 kg, respectively for farm 1 and 2. For Jamunapari goat's kid birth weight were 1.51 +/- 0.07 and 1.42 +/- 0.09 kg, for male and females, respectively in the farm 2. The average weaning age was 4 months and the average weaning weight of male and female black Bengal goats were 5.19 +/- 0.358, 5.05 +/- 0.28, 5.63 +/- 0.61 and 5.54 +/- 0.41 kg, in the farm 1 and 2, respectively. However, the average weaning weight of male and female Jamunapari was 6.59 +/- 0.69 and 6.79 +/- 0.31 kg, respectively in farm 2 which was higher than black Bengal. The average age at sexual maturity of black Bengal goat was 8 months. The average weight at sexual maturity of male and female black Bengal goats were 9.82 +/- 0.75 and 9.52 +/- 0.62 kg, respectively in farm 1 and 9.65 +/- 0.75 and 9.138 +/- 0.70 kg, respectively in farm 2. The average age at sexual maturity was 9 months for Jamunapari goat. The average weight at sexual maturity of male and female Jamunapari goats was 13.2 +/- 0.75 and 14.1 +/- 0.82 kg, respectively. The average daily body weight gain from birth to weaning for male and female black Bengal goat was 33.70, 35.11 g day(-1) and was 35.67 and 45.94 g day(-1), respectively in farm 1 and 2 and for Jamunapari goat was 42.97 and 45.47 g day(-1), respectively. The males were grew faster than the females. The predicted live weight gains for both breeds were lowered after fitting the linear regression. The co-efficient of determination (R(2)) of male and female black Bengal and Jamunapari goats kids similar.

Thalassaemia: a study on the perception of patients and family members.

Marked improvement in the management of thalassaemia has not been matched by progress in psychosocial rehabilitation as thalassaemia continues to pose challenges to patients and their family members. Few studies have been carried out in Malaysia to look at such issues. This study is therefore to explore the concerns, beliefs and feelings about thalassaemia. It was conducted in the year 2009 over 7 months on "focus groups", in patients aged 8-22 years and parents attending Paediatric Clinic of Tengku Ampuan Afzan Hospital, Kuantan, Pahang. Results showed that concerns and adverse impact were related to lower grades in education, poor self-image, less chance of employment, marriage, financial burden and social integration. Compliance to subcutaneous iron chelator was poor. There were various concerns related to blood transfusion therapy. It is evident that thalassaemia greatly affects the psychosocial dimensions and a more structured long term psychosocial support is needed to improve quality of life of patients.

Screening for thalassaemia among group of students of a higher institution – our experience

Thalassaemia is an inherited blood disorder and is a significant public health alarm in Malaysia with many not knowing they are carriers of this haemoglobin disorders. Materials and methods: This study conducted a one off collection of blood samples from 72 Malays students of International Islamic University Malaysia (IIUM) in Kuantan. Blood samples were subjected to conventional haemoglobin analyses that include full blood count and picture, HPLC, Haemoglobin electrophoresis and H-inclusion test. All samples were also genotyped for alpha thalassaemia–1 of Southeast Asia (a-Thal1SEA). Result: There were 17(23.6%) students who were diagnosed as thalassaemia carriers. Out of this, four (5.5 %) and six (8.3 %) students were presumptive β-thalassaemia trait and Haemoglobin-E trait as determined by the HPLC assay respectively. Nine (12.5%) students were genotyped a-Thal1SEA among whom two were also β-thalassaemia carriers. All thalassaemia cases had MCH of 80fL. Two out of four (50%) presumptive β -thalassaemia trait and one out of six (17%) students of presumptive Haemoglobin-E trait had family history of thalassaemia respectively. Conclusion: The high occurrence of the three common types of thalassaemia carrier (β, Hb-E and a-Thal1SEA thalassaemia) in our small group of subjects could be due to better participation of students who had family history of thalassaemia. The study reaffirmed the importance of molecular study for detection of alpha-thalassaemia and the use of MCH value of <27pg rather than MCV value of < 80fL for prediction of thalassaemia.

Rescue of renal function in a 3-year-old girl with Goodpasture's syndrome with a brief review of literature.

Goodpasture's syndrome has been documented in only a handful of children under the age of four. We describe a 3-year-old girl presenting with anaemia and renal failure whose kidney biopsy showed anti-glomerular basement membrane (GBM) disease. She was treated aggressively with pulse steroids, plasmapheresis and monthly infusions of cyclophosphamide. After months of aggressive immunosuppression, her renal function normalized, and her anti-GBM antibody disappeared. A year after the onset, she underwent a second kidney biopsy for persistent proteinuria and hypertension that surprisingly showed focal sclerosing glomerulonephritis, an unreported finding at this age. The biopsy showed deposition of antibody on the GBM despite the fact that anti-GBM antibody had normalized in the serum 5 months earlier. Mycophenolate mofetil was added to the immunosuppression at that point. At her 3-year follow-up, creatinine clearance was 88.4 mL/min/1.73 m(2), proteinuria was 408 mg/day and blood pressure was controlled with enalapril 0.2 mg/kg/day. She has not had a relapse or abnormal anti-GBM antibody for 30 months, but her renal prognosis remains guarded. To our knowledge, this is the youngest patient to have a successful rescue of renal function after isolated Goodpasture's syndrome.

Non-steroidal anti-inflammatory drugs-associated acute interstitial nephritis with granular tubular basement membrane deposits.

Acute tubulo-interstitial nephritis (ATIN) is an important cause of acute renal failure resulting from a variety of insults, including immune complex-mediated tubulo-interstitial injury, but drugs such as non-steroidal anti-inflammatory drugs (NSAIDs) are a far more frequent cause. Overall, as an entity, ATIN remains under-diagnosed, as symptoms resolve spontaneously if the medication is stopped. We report on a 14-year-old boy who developed acute renal failure 2 weeks after aortic valve surgery. He was put on aspirin following surgery and took ibuprofen for fever for nearly a week prior to presentation. He then presented to the emergency department feeling quite ill and was found to have a blood urea nitrogen (BUN) concentration of of 147 mg/dl, creatinine of 15.3 mg/dl and serum potassium of 8.7 mEq/l. Dialysis was immediately initiated. A kidney biopsy showed inflammatory infiltrate consistent with ATIN. However, in the tubular basement membrane (TBM), very intense granular deposits of polyclonal IgG and C3 were noted. He needed dialysis for 2 weeks and was treated successfully with steroids for 6 months. His renal recovery and disappearance of proteinuria took a year. In conclusion, this is a first report of NSAIDs-associated ATIN, showing deposits of granular immune complex present only in the TBM and not in the glomeruli.

Favourable response to splenectomy in familial myelodysplastic syndrome.

Familial myelodysplastic syndrome occurring at a young age is a very rare childhood haematological malignancy. Two siblings, aged three and 18 years, from a consanguineous marriage, presented with pancytopenia and was subsequently diagnosed to have myelodysplastic syndrome. Both remained clinically stable throughout the illness. Splenectomy appeared to have fully corrected the cytopenia in one of them.

Effects of angiotensin II on NaPi-IIa co-transporter expression and activity in rat renal cortex.

The kidney plays a major role in reabsorption of phosphate with the majority occurring in the proximal tubule (PT). The type IIa sodium-phosphate co-transporter (NaPi-IIa) is the main player in PT. The purpose of current study was to determine the effect of angiotensin II (A-II) on membrane expression of NaPi-IIa in the rat renal cortex. A-II (500 ng/kg/min) was chronically infused into the Sprague-Dawley rats by miniosmotic pump for 7 days. The arterial pressure and circulating plasma A-II level along with urine output were markedly increased in A-II rats. There was diuresis but no natriuresis. The phosphate excretion increased sevenfold on day 4 and 5.7-fold on day 7. There was no change in Na-dependent Pi uptake in brush-border membrane (BBM) vesicles between A-II-treated group and control on day 4, however, there was a 43% increase on day 7. Western blot analysis of NaPi-IIa protein abundance showed a parallel pattern: no change after 4 days of treatment and a 48% increase after 7 days of treatment. However, Northern blot analysis of cortical RNA showed no change in NaPi-IIa mRNA abundance on day 7. A-II stimulation of Na/Pi co-transport activity is a result of increases in the expression of BBM NaPi-IIa protein level and that stimulation is most likely mediated by posttranscriptional mechanisms.

Kimura disease with advanced renal damage with anti-tubular basement membrane antibody.

Kimura disease (KD) is an autoimmune eosinophilic granulomatous disorder with generalized lymphadenopathy. A handful of pediatric patients with renal disease have been described, none of whom have been African-American (AA). We present an AA boy with KD and nephrotic syndrome (NS). Two months after stopping steroids, fever, asthma, eczema, and proteinuria recurred. His NS did not relapse but his platelet count decreased to 51,000/microl (x10(6)/l). On restarting prednisone, his platelet count normalized. A kidney biopsy revealed 23 of 37 glomeruli obsolescent and advanced damage with over 50% of cortical tissue replaced by interstitial fibrosis and chronic inflammation. Glomerular immunofluorescence was largely negative; very intense linear anti-tubular basement membrane (TBM) deposits of IgA, IgG, C3, and C4 were noted. At present, 36 months from onset, serum creatinine is 1.2 mg/dl (106 micromol/l). We present a 4-year-old AA boy with KD, NS, relapsing thrombocytopenia, and renal damage with anti-TBM antibody.

Transient hypothyroidism in a breastfed infant after maternal use of iodoform gauze.

We describe transient hypothyroidism induced in a 2 week-old breastfed infant by maternal use of iodoform gauze for a perirectal abscess. The initial newborn thyroid screen was normal. Low levels of thyroid hormone were discovered during a routine second newborn screen. Thyroid function has since normalized with short-term therapy with thyroxine and after the mother discontinued use of the packing iodoform gauze. This case illustrates the need to avoid iodine-containing products in breastfeeding mothers.

Hyponatremic hypertensive syndrome (HHS) in an 18-month old-child presenting as malignant hypertension: a case report.

BACKGROUND: The combination of hyponatremia and renovascular hypertension is called hyponatremic hypertensive syndrome (HHS). Malignant hypertension as a presentation has been reported in adults with HHS but is rare in children. CASE PRESENTATION: An eighteen month-old male presented with drowsiness, sudden onset status epilepticus and blood pressure of 210/160. The electrolytes on admission revealed sodium of 120 mEq/L and potassium of 2.1 mEq/L. The peripheral renin activity (PRA) was 172 ng/ml/min (normal 3-11 ng/ml/min) and serum aldosterone level was 91 ng/dl (normal 4 to 16 ng/dl). Patient underwent angioplasty with no success, followed by surgical correction. Two years since the diagnosis, the blood pressure is controlled with labetolol and amlodipine (at less than sixth of the pre-operative dosages). The PRA is 2.4 ng/ml/min and aldosterone 15.5 ng/dl. The child not only had three renal arteries on left but all of them were stenosed which to best of our knowledge has not been described. CONCLUSION: As uncommon as HHS with malignant hypertension may be in adults it is under-reported in children and purpose of the case report is to raise its awareness.

Managing Henoch-Schonlein purpura in children with fish oil and ACE inhibitor therapy.

BACKGROUND: Henoch-Schonlein purpura (HSP) is a vasculitic syndrome with palpable purpura and renal involvement. The treatment for HSP with persistent renal disease remains controversial. The kidney biopsy in HSP shows IgA deposits and fish-oil therapy has proven to be promising in halting the progression of IgA nephropathy. METHODS: Five children with biopsy-proven HSP with repeated episodes of haematuria and proteinuria were treated with fish oil (1 g orally twice daily). In three of the five patients an angiotensin-converting enzyme inhibitor (ACEI) was added for hypertension. RESULTS: The mean duration of follow up after starting fish-oil therapy was 49.2 weeks. The protein excretion rate prior to starting fish oil was 1041 mg/day and on the last follow-up visit the rate had decreased to 104 mg/day (P <0.05). The average blood pressure (BP) prior to therapy was 135/82. On the last follow-up visit the average BP off ACEI had decreased to 100/54 (P <0.05). After a year of follow up serum creatinine and glomerular filtration rates have remained stable at 51.2 micromol/L and 128 mL/min/1.73 m2, respectively. CONCLUSION: This is the first report of abatement of HSP with fish oil and ACEI in children. There is a need for randomized prospective trials to confirm this observation.