The repair process of airway epithelium involves cell migration, spreading, proliferation and re-differentiation. Objective: Cellular and molecular responses to tracheal brush induced injury were investigated using a rabbit model. Methods: Eighteen New Zealand white rabbits were divided into uninjured and injured groups. After tracheal brushing, the animals were maintained in the laboratory before being sacrificed at given time points (1, 12 hours, 3, 7, and 21 days). The trachea of each rabbit was retrieved and preserved before being subjected to haematoxylin and eosin staining and real time PCR. Results: After injury, the remained epithelial cells underwent an instant response by proliferating and migrating into the damaged site. This finding was in accordance with the proliferative and migration activity-related gene expression results (MMP-9, TIMP1, vimentin, and ß-integrin). The increased activity of these genes was crucial at the early time points, as it encouraged the remaining cells to repopulate the damaged area. Conclusions: Continuous regulation of MMP-9, vimentin and ß-integrin plays important roles in promoting cellular homing especially the cells bordering the lesion to migrate and repair of the damaged ECM. Thus, this activation enhanced regeneration and repair of the damaged tracheal epithelium as early as 1 h and complete at 21 d following injury.
Immunization has been introduced for decades to eradicate fatal infectious diseases by inoculating attenuated, killed or toxoid of microorganisms such as bacteria and virus. The triggering action to the immune system would not harm the host; despite can boost the immune responses to any infection. However, several cases of the eradicated infectious disease have re-emerged due to the existence of vaccine hesitancy group. Vaccine hesitancy has been observed emerging worldwide due to rejection in receiving vaccine. The main obstacle in vaccination program was identified according to the misconception that they received from internet or any mass media without boundaries. Various actions from the government have met the needs to enforce and educate the public especially the hesitant group towards better disease prevention with vaccination. The strategy would cover any interaction activities or programs with the public in transferring the information about the vaccination and its benefit to the health of herd community.
Introduction: Methylation of promoter region of p16 leading to gene silencing has been implicated ina wide range of malignancies including lymphomas. In diffuse large B cell lymphoma (DLBCL) particularly, a varying percentage of epigenetic inactivation of p16 promoter region was observed ranging from 16 -54%. However, quantitative analysis of p16 promoter methylation in DLBCL has not been extensively studied in Malaysia. Objective: This study aims to quantitatively analyse p16 methylation in DLBCL samples using pyrosequencing technique. Methods: Genomic DNA was extracted from 16 formalin-fixed paraffin-embedded lymphoma tissue blocks from patients diagnosed with DLBCL. Samples were retrieved from Hospital Tengku Ampuan Afzan, Pahang and Hospital Universiti Sains Malaysia, Kelantan. Primers were designed to amplify bisulfite-treated DNA targeting p16 promoter region. Methylation status of 7 CpG sites was determined by pyrosequencing. Results: All the 16 samples studied showed promoter methylation of p16. The range of mean methylation percentage was between 18 to 81%. Conclusion: The present study has successfully measured the level of methylation of p16 in all 7 CpG sites despite the limitation in sample size. Since p16 methylation is a common event in our series of DLBCL cases, it is worth including a larger sample size in future studies to increase the chance of finding a significant correlation with clinical parameters.
Acute lymphoblastic leukemia (ALL) is the most common
childhood leukaemia. It is a malignant neoplasm caused by the proliferation of
poorly differentiated precursors of the lymphoid cells. It is relatively
uncommon in adult. In adult ALL, central nervous system (CNS) involvement
is associated with poor prognosis. The incidence of CNS involvement has
been reported between 7% and 15 %. We report a case of optic nerve
infiltration in ALL in a 49 years old gentleman. He was diagnosed with
precursor-B ALL. He was treated with chemotherapy and CNS prophylactic
regime. He presented with sudden left eye loss of vision for one-day duration
with history of right eye inferior visual field loss for the past three months. His
visual acuity was no perception to light on the left eye and 6/9 on the right
eye. There was marked left relative afferent pupillary defect. The right eye
showed decreased in optic nerve function with inferior visual field defect.
Anterior segment examination was unremarkable in both eyes. Left optic disc
appeared normal but the right optic disc was pale. Blood investigation
showed no sign of infection or haemoconcentration. Cerebral spinal fluids
examination revealed abundant of white cells and blast cells. Magnetic
resonant imaging showed bilateral optic nerve enhancement suggesting of
bilateral optic nerves infiltration. He was started on a new regime of
chemotherapy followed by cranial radiotherapy. Unfortunately, he succumbed
to death due to septicaemia. There are variations in clinical presentation of
optic nerve infiltration in leukaemic patients. Normal appearance of optic disc
may not exclude the possibility of infiltration by malignancy. Assessment of
the optic nerve function and imaging is helpful for the detection of leukaemic
infiltration. Early detection of optic nerve infiltration is important for initiation or
change of therapy to prevent mortality.
Introduction: Isolation of specific cell types is important in providing a better understanding of hematological disorders. The knowledge of molecular biology aspect in β-thalassemia is still limited. This is because hemoglobin disorder involves various erythropoietic processes in which the genetic information is lack due to enucleation of red blood cells occurs in bone marrow. It is invasive to collect samples from bone marrow and cord blood although nucleated red blood cells (NRBCs) are abundant in these sites. NRBCs are precursors of red blood cells and typically found in peripheral blood (PB) of β-thalassemia major patients and abundant post-splenectomy. The utilization of PB NRBCs will provide a further understanding of the molecular aspects of ineffective erythropoiesis in β-thalassemia major patients. Objective: The objective of this study was to isolate the NRBCs using CD71 magnetic beads from PB of β-thalassemia major; non-splenectomy and post-splenectomy patients. Methods: NRBCs were isolated from 6 mL PB of β-thalassemia major patients based on density gradient and magnetic activated cell sorting (MACS) for NRBCs enrichment using a CD71 marker. Cell count was determined by using hemocytometer (Weber Scientific, NJ, USA) and BD FACSCantoTM II flow cytometry (Becton-Dickson, NJ, USA) was performed for method validation. Results: NRBCs were successfully isolated from the PB of both non-splenectomy and post-splenectomy β-thalassemia major patients with >90% specificity by flow cytometric analysis. The median number of enriched NRBCs (x104 ) was 58.5 (283) and 340 (338) respectively using hemocytometer. Conclusion: The MACS method was found to be convenient and efficient in the isolation of the targeted cells for downstream applications.
Mutations in the δ globin gene are not pathologically significant [1]. However, coinheritance of β and δ thalassaemia can mask the diagnosis of β thalassaemia trait as it causes HbA2 level to be lowered [2,3]. Here, we reported 5 unrelated cases of compound heterozygous β0 Filipino ~ 45 kb deletion and codon 67 (GTG>ATG) HbA2 Deventer in Sabahan population.
Cases of β°-thalassemia traits with unusual low HbA2 were reviewed. These cases were initially referred to our laboratory for definitive diagnosis of β-thalassemia trait. Haematological parameters and Hb analysis were carried out at the referral hospital. Genomic DNA was extracted from the peripheral blood. Multiplex ARMS and Gap PCR were done to detect common point mutations and deletions for both alpha and beta globin genes. Sanger sequencing was performed to detect mutations in delta globin gene.
Patients’ consist of 4 males and 1 female aged between 25-38 years old. All of them are indigenous Sabahan (2 Kadazans, 1 Murut, 1 Dusun and 1 Sungai). Their haemoglobin level ranges between 10.8 – 12.8g/dl. Hb analysis findings of HbA2 and HbF level ranges between 2.9 – 4.0 and 2.2 – 9.4g/dl respectively. Molecular findings revealed heterozygous state of (β)º-thal, Filipino ~45Kb deletion, NG_000007.3:g.[66258_184734del];[66258_184734=] and heterozygous state of Codon 67 [GTG>ATG] Hb A2-Deventer mutation, NG_000007.3:g.[63512G>A];[63512G=] (Figure 1 and 2).
Detection of 5 unrelated cases of HbA2 Deventer may suggest that this delta variant is common among indigenous Sabahan. Since beta thalassaemia is also common in the population, more attention should be paid during diagnosis. Identification of delta variant in beta thalassaemia carrier is important because coinheritance of beta and delta thalassaemia results in a less elevated HbA2 level. Therefore, molecular testing of thalassemia carrier state in the case of borderline HbA2 is warranted to avoid misdiagnosis of beta thalassaemia carriers.
Chromosomal abnormalities (CA) can affect numerical or structural compositions of chromosomosal DNA leading to a diversity of clinical phenotypic presentations. Awareness of prenatal diagnosis and genetic counselling have improved with advancing medical research but CA remain prevalent as its aetiology is unknown. The objective of this study is to determine the frequencies of various CA in the principle region of north-western Malaysia and compare this data to previous reports to ascertain if statistical differences exist. Karyotype analyses performed at the Genetics Laboratory, Advanced Diagnostic Laboratory (ADL) during the first 5-years of cytogenetic services, totalling 1461 cases, were assessed in this report. Cases suspected of CA were initially diagnosed by clinicians and detailed clinical and family histories were recorded. Peripheral blood lymphocytes of patients were collected and cultured in vitro for acquisition of karyotype by standardized G-banding technique. Fluorescence in situ hybridization (FISH) was conducted in cases suspected of to be DiGeorge, Prader-Willi, Angelman and Williams syndrome. Of the total samples (1805) received and cultured, 1669 (92.46%) successfully yielded results. Abnormal outcomes were observed in 495 cases (29.66%) whereby pronounced majority of cases 299 (68.42%) were Down syndrome. This is followed by Edward, Turner and Patau syndrome, in order of frequency. Numerical CA appears to be prevalent accounting for 85.86% of cases. Structural CA accounted for 14.14% of total positive cases whereby the most common was deletions (34.29%) followed by translocations (20%), ring chromosomes (5.71%), Fragile X syndrome (4.29%), duplications (5.71%) and marker chromosomes (7.14%). The remainder of cases (22.86%) consisted of derivative chromosomes and other complex aberrations. The number of polymorphic variant cases were 27 (1.62%). The number of peripheral blood samples received has significantly increased from 14.3 per month in 2006 to 32.17 per month in 2011. Comparative analysis of our study to previous reports reveal statistical differences in the occurrence of several CA including Edward, Patau, Klinefelter and Fragile-X syndrome. Our experience with peripheral blood samples for cytogenetic analysis demonstrated a success rate of 92.46%. This showed an increase in clinicians validating patients’ diagnoses with karyotyping which is essential in confirming genetic anomalies with the goal to substantiate genetic counselling.
The most common inherited monogenic disorders in the world are the haemoglobinopathies and thalassaemia. Thalassaemia is a heterogeneous group of genetic disorders of haemoglobin synthesis, characterised by a reduction in the production of one or more of the subunits of haemoglobin chains [1]. Haemoglobin A2 (HbA2) level is an important parameter in thalassaemia diagnosis. High HbA2 level (≥4.0) detected in Hb analysis, points to the diagnosis of beta thalassaemia and other haemoglobinopathies. However, in some cases, the HbA2 levels are apparently normal or borderline high despite abnormal haematological profile. In these cases, further testing is required to confirm the diagnosis. The aim of this study is to examine any abnormality at molecular level in cases of Hb analysis results with normal or borderline high HbA2 level.
The aims of this study are to identify and characterize the Haemoglobin G Makassar. Haemoglobin G Makassar was identified in Makassar, Sulawesi (Celebes), Indonesia in 1969 and has been reported in a family of Thai origin in 2002. Haemoglobin G Makassar was found to share identical properties with haemoglobin S in routine haemoglobin separation by cation-exchange HPLC. It is therefore, patients with Haemoglobin G Makassar and Haemoglobin S may sometimes be mistakenly identified for each other.
There were four cases identified from year 2015 to 2016 in Peninsular Malaysia by Molecular Genetics Laboratory, Institute for Medical Research. All patients were asymptomatic with mild hypochromic microcytic anaemia. All patients were analysis with Haemoglobin S trait. Analysis by Capillary Electrophoresis showed that these patients had 39.9 to 44.0% of haemoglobin variant in zone S. Alpha and Beta globin gene analysis were performed on these samples.
DNA sequence analysis, revealed a single nucleotide substitution GAG to GCG at codon 6 of the beta-globin gene (Glu>Ala), indicating of Haemoglobin G Makassar for all the patients (Fig. 1). All patients were positive with Haemoglobin S trait. Multiple Amplification Refractory Mutation System (MARMS) PCR for Haemoglobin S was negative in all cases. However alpha-globin gene analysis showed that two of them had single alpha deletion (α3.7). The mean reading for HGB is 11.95 g/dL, for MCV is 72.1 fL and for MCH is 23.65 pg which all are lower than normal peoples.
The screening method may mistakenly identify Haemoglobin G Makassar as Haemoglobin S. Therefore identification and characterization of Haemoglobin G Makassar by several molecular methods such as polymerase chain reaction (PCR) and sequence analysis are necessary for confirmation of the diagnosis.
Macular branch retinal vein occlusion (BRVO), a type of retinal vein occlusion, is rarely recognised as a distinct entity. Macular BRVO has unique clinical features and different natural courses than the major BRVO. We report a case of a young patient with macular BRVO with macular oedema who was successfully treated with intravitreal ranibizumab injection. A 43 year-old Chinese man with no underlying medical illness presented with 2 weeks history of left eye painless reduced central vision which was worsening over time. On examination, his left eye visual acuity was 6/30 and Amsler chart drawing showed a lower central scotoma. Dilated fundus examination found marked flame-shaped retinal hemorrhages with cotton wool spot over the superior macular area bounded superiorly by superior arcade and macular thickening. An optical coherence tomography revealed cystoid macular oedema; and fundus fluorescein angiography showed occlusion of a small venous branch draining a superior part of macula to superior temporal venous arcade. A complete medical investigation found that he has hypertriglyceridemia and he was managed accordingly. His vision had improved to 6/6 after receiving 3 injections of intravitreal ranibizumab with no residual central scotoma and complete resolution of macular oedema.
The presence of retinal vasculitis in patients with pulmonary tuberculosis is not uncommon. However, asymptomatic presentations are quite rare. Here, we present a case of 25-year-old Malay gentleman with pulmonary tuberculosis, who was referred for an eye assessment following initiation of anti-tuberculosis drugs. His vision was good; he had no other symptoms despite having retinal neovascularization, pre-retinal haemorrhages and healed choroiditis at the fundus of the left eye. Fundus fluorescein angiography showed multiple areas of capillary fallouts at the superotemporal quadrant of the left eye with leakage from the retinal neovascularization. He was treated with laser retinal photocoagulation and oral steroids for 6 weeks, while the anti-tuberculosis drugs were continued for 9 months. Subsequent follow-ups showed regression of the neovascularization. He has remained asymptomatic since his initial visit to the eye clinic.