Green tea polyphenol "epigallocatechin-3-gallate", differentially induces apoptosis in CLL B-and T-Cells but not in healthy B-and T-Cells in a dose dependant manner.

B-cell chronic lymphocytic leukaemia (CLL) is characterized by an accumulation of CD5-positive monoclonal B-cells due in large part to a failure of apoptosis. The ability to study CLL B-cells in vitro has always been a challenge and hampered by the low viability of the CLL B-cells in cell culture systems. In this study, we present a multicellular cell culture system to maintain CLL B-cells viable in culture for 60h in the presence of a stromal cell feeder layer in combination with a whole white blood cell preparation.

RNF187 is Downregulated Following NF-κB Inhibition in Late Erythroblasts.

Beta (β)-thalassaemic erythroblasts grown in vitro have reduced nuclear factor kappa B (NF-κB) pathway gene expression. By inhibiting this pathway in erythroblasts from normal individuals, important downstream genes affected by this inhibition can be identified. Bay 11-7082 is a potent inhibitor of the NF-κB pathway, it acts irreversibly, inhibiting NF-κB activation by blocking tumor necrosis factor alpha (TNF-α)-induced phosphorylation of the inhibitory IκB subunit thereby preventing NF-κB activation.

The Effect of Nonsense Mediated Decay on Transcriptional Activity Within the Novel β-Thalassemia Mutation HBB: c.129delT.

Premature termination codons (PTCs) are caused by mutations in the coding sequences of functional genes resulting in an incorrect assignment of a stop codon. Abnormal and truncated proteins are prevented from being translated due to the rapid degradation of mRNA carrying these mutations by an RNA surveillance mechanism referred to as nonsense mediated decay (NMD). Recently, a novel mutation in a patient from Thailand with the clinical diagnosis of Hb E (HBB: c.

Experimental Characterization of Hb Flurlingen (HBA2: c.177 C > G, p.His > Gln) and Hb Boghé (HBA2: c.177 C > A, p.His > Gln) Reveals Contradictory HBA2 Expression and Translation Patterns Despite Identical Amino Acid Substitutions.

In this study, we describe the clinical features and provide experimental analyses of Hb Flurlingen (HBA2: c.177 C > G, p.His > Gln) that contrasted with Hb Boghé (HBA2: c.

Differential gene expression analysis in early and late erythroid progenitor cells in β-thalassaemia.

β- thalassaemia is a disorder of globin gene synthesis resulting in reduced or absent production of the β-globin chain in red blood cells. In this study, haematopoietic stem cells were isolated from the peripheral blood of six transfusion dependent β-thalassaemia patients and six healthy controls. Following 7 and 14 d in culture, early- and late- erythroblasts were isolated and purified.

Molecular characterization of Hb Hamilton Hill (HBA2: c.388delC), a novel HBA2 variant generating a premature termination codon and truncated HBA2 chain.

In recent years, the identification of α-thalassemias caused by nondeletional mutations has increased significantly due to the advancement of sensitive molecular genetics tools. We report clinical and experimental data for a novel frameshift mutation caused by a single base deletion at position 388 in exon 3 of the α2-globin gene (HBA2: c.388delC; Hb Hamilton Hill), resulting in the phenotype of α-thalassemia (α-thal).

Molecular and cellular analysis of three novel alpha2-globin gene promoter mutations [HBA2: c.-59C>T], [HBA2: c.-81C>A] and [HBA2: c.-91G>A] reveal varying patterns of transcriptional and translational activities.

While point mutations affecting the promoter region of β-globin gene are widely described, there are no well characterised reports of any point mutations currently found in the promoter of the α2-globin (HBA2) gene. We present clinical and experimental data for three novel HBA2 gene core and proximal promoter mutations. Using an in vitro system designed to assess the impact of point mutations, the three novel [HBA2:c.

Molecular and cellular analysis of a novel HBA2 mutation (HBA2: c.94A > G) shows activation of a cryptic splice site and generation of a premature termination codon.

In this study, we describe the clinical features and provide experimental analyses of a novel point mutation affecting the penultimate nucleotide of the first exon of the HBA2 (HBA2: c.94A > G) gene identified in a 26-year-old female who also carries a heterozygous Hb E (HBB: c.79G > A) variant.

Alpha thalassaemia due to non-deletional mutations on the -3.7 alpha globin fusion gene: laboratory diagnosis and clinical importance.

Aims: Alpha (α) thalassaemia may be caused by large deletions of the α globin gene(s), or rarely, non-deletional mutations. Both types of mutations may co-exist, and if located on the same allele (α), produce a reproductive risk of hydrops fetalis. We illustrate how clinical-laboratory correlation and accurate α gene sequencing are essential in identifying such patients.

Differential expression of the Bcl-2 and Bax isoforms in CD19 positive B-lymphocytes isolated from patients diagnosed with chronic lymphocytic leukaemia.

Aim: To examine the relative gene expression levels of the anti-apoptotic Bcl-2α and β isoforms and the pro-apoptotic Baxα and β isoforms in patients with chronic lymphocytic leukaemia (CLL) and healthy controls (HC).

Methods: Peripheral blood was obtained from 36 patients diagnosed with CLL and 10 HC. CD19 B-lymphocytes were isolated using an antibody coupled magnetic bead isolation system; from these cells the total RNA was isolated and purified.

α-thalassemia trait caused by frameshift mutations in exon 2 of the α2-globin gene: HBA2:c.131delT and HBA2:c.143delA.

We describe two frameshift mutations associated with an α-thalassemia (α-thal) phenotype, identified in three unrelated individuals investigated for persistent microcytosis. The first mutation, HBA2:c.131delT, is located in codon 43, and the second, HBA2:c.

Identification and characterization of two novel and differentially expressed isoforms of human α2- and α1-globin genes.

In most references, the transcription initiation site for the α2- and α1-globin genes has been described to lie 37 bp upstream of the translation initiation codon, however, a review of data repositories such as GenBank and Ensembl showed a report of the α2-globin transcription initiation site occurring at position -66 relative to the initiation codon. To confirm the occurrence of these isoforms for both the α2- and α1-globin genes and to document their expression levels, we initiated our current investigation. Total RNA from the peripheral blood of 15 healthy volunteers was analyzed using both semi-quantitative-polymerase chain reaction (PCR) and real-time (ReTi-PCR) protocols developed in our laboratory, with primers designed to enable distinction between the α2- and α1-globin transcripts.

A molecular tool to assess the pathological relevance of alpha-globin DNA variants.

Aim: While the phenotype for heterozygous beta-thalassaemia is straightforward, it is more difficult to confirm a causative relationship for mutations in the alpha-globin genes. The aim of this study was to generate an in vitro system to evaluate the pathological relevance of α-globin mutations.

Methods: The novel variant HBA1:c.

Molecular and cellular characterization of a new α-thalassemia mutation (HBA2:c.94A>C) generating an alternative splice site and a premature stop codon.

The identification of α-thalassemia (α-thal) due to point mutations has been increasing significantly with the advancement of molecular diagnostic tools. We describe here the molecular and cellular characteristics of the thalassemia mutation HBA2:c.94A>C, a novel point mutation affecting the α2-globin gene, causing a mild α-thal phenotype in a male patient of undisclosed ethnicity, investigated for unexplained microcytosis.

In vitro characterization of the α-thalassemia point mutation HBA2:c.95+1G>A [IVS-I-1(G>A) (α2)].

The α-thalassemias are a group of disorders occurring as a result of decreased synthesis of α-globin chains, most commonly due to deletions of α-globin genes. Detection of α-thalassemia (α-thal) caused by point mutations has increased during the past few years and more than 70 different point mutations have been reported for the α1- and α2-globin genes. The mutation at the splice donor site of the first intervening sequence [IVS-I-1 (G>A)] of the α2-globin gene, HBA2:c.

Tympanic membrane wound healing in rats assessed by transcriptome profiling.

Objectives/hypothesis: The aim of this study is to elucidate transcriptional changes that occur in response to tympanic membrane (TM) perforation in rats and to infer key genes and molecular events in the healing process.

Study Design: A prospective cohort study of 393 male Sprague-Dawley (Rattus norvegicus) rats.

Methods: Sprague-Dawley rats were randomly allocated into either control or perforation groups spanning a 7-day time period.

Incidence of c-Cbl mutations in human acute myeloid leukaemias in an Australian patient cohort.

Aim: The aim of this study was to investigate the incidence and characteristics of c-Cbl mutations in acute myeloid leukaemias (AMLs) from an Australian patient cohort. Two initial studies examining c-Cbl mutations in AML, one from Germany and one from the US, found vastly different incidences of mutations (0.6% compared to 33%, respectively).

Hb Lynwood [α107(G14) (-T) (α2) HBA2:c.323delT)] in conjunction with the α(3.7) deletion produces a moderately severe α-thalassemia phenotype.

We describe a novel frameshift mutation associated with an α-thalassemia (α-thal) phenotype in a patient of Sudanese origin investigated for persistent microcytosis. In addition to the α(3.7) deletion, a novel mutation on the α2 gene was detected: HBA2:c.

Keratinocyte growth factor 1, fibroblast growth factor 2 and 10 in the healing tympanic membrane following perforation in rats.

The aim of this study was to provide a transcriptome profile of Keratinocyte Growth Factor (KGF)-1, Fibroblast Growth Factor (FGF) 2 and FGF10 (KGF2) in the healing rat tympanic membrane (TM) over 7 days and an immunohistochemical account over 14 days following perforation. KGF1, FGF2, and FGF10 play important roles in TM wound healing. The tympanic membranes of rats were perforated and sacrificed at time points over a 14-day period following perforation.

Hb East Timor [β80(EF4)Asn→His, AAC>CAC (HBB c.241A>C)], a variant hemoglobin associated with normal hematology.

Routine hemoglobin (Hb) analyses identified a new β-globin variant in a family from East Timor. The red cell indices were within normal limits for all affected family members. The variant is due to a missense mutation at amino acid codon 80 (AAC>CAC) which results in the substitution of histidine for asparagine.

The role of epidermal growth factor in the healing tympanic membrane following perforation in rats.

Epidermal Growth Factor (EGF) has been identified as playing a critical role in the wound healing process. The objective of this study is to investigate the role that EGF plays in rat tympanic membrane (TM) wound healing using two techniques, microarray and immunohistochemistry. The tympanic membranes of rats were perforated using a sterile needle and sacrificed at time points during 2 weeks following perforation.

Histology of the healing tympanic membrane following perforation in rats.

Objectives/hypothesis: The aim of this study was to provide a detailed cytological account on the healing tympanic membrane (TM) over 14 days and to complement existing research into TM wound healing.

Study Design: The study is a prospective cohort study of 19 male Sprague-Dawley (Rattus norvegicus) rats.

Methods: Rat TMs were perforated using a sterile needle and sacrificed at time points during the 14 days following perforation.

Advancing towards a tissue-engineered tympanic membrane: silk fibroin as a substratum for growing human eardrum keratinocytes.

Human tympanic membrane cells (hTMCs), harvested from tympanic membrane (TM) explants, were grown in culture and then seeded on membranes prepared from silkworm (Bombyx mori) silk fibroin (BMSF) and on tissue-culture plastic membranes (PET). Fibroin was isolated from silk cast into membranes with a thickness of 10-15 microm. The hTMCs were cultured on both materials for 15 days in a serum-containing culture medium.

Chronic tympanic membrane perforation: a better animal model is needed.

Developments in the treatment of chronic tympanic membrane perforation have been hindered by the lack of an ideal animal model. It is not appropriate to test such treatments on acute perforations as the majority of these heal spontaneously. An ideal animal model would be one that most closely resembles the human clinical situation.