Molecular gene cloning and sequencing of glutamate decarboxylase gene from Lactobacillus delbrueckii and Lactobacillus reuteri

Glutamate decarboxylase enzyme produces ?-aminobutyric acid [GABA] in a non-reversible decarboxylation reaction of glutamate. GABA is a major inhibitory neurotransmitter of the brain and it is also present at high concentration in other organs such as pancreatic islets. GABA has effects on blood pressure, diabetes, inflammation, sleeplessness and depression. Some bacteria such as Lactobacillus strains are capable of GABA production. Identification of these bacteria is important both for researchers and industry. The aim of this study was molecular gene cloning and sequencing of glutamate decarboxylase [gad] from Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 and Lactobacillus reuteri ATCC 23272. These strains were cultured in MRS medium at 37degreeC for 24 hours. For cloning gad gene from these strains, polymerase chain reaction [PCR] was performed using specific primers designed by Oligo7 software. PCR production was extracted from agarose gel and was inserted into PGEM-T vector using T4 DNA ligase enzyme and then it was transformed to E. coli XL1Blue. In the final step, white colonies were selected and after plasmid extraction, the existence of gad gene in recombinants was confirmed by PCR. Gad gene was cloned from Lactobacillus delbrueckii subsp. bulgaricus ATCC 11842 and Lactobacillus reuteri ATCC 23272. It is for the first time that the gad gene sequences from these bacteria were registered on NCBI with accession numbers KF751355 and KF751352 respectively. The result of this research indicates that the two aforementioned bacteria contain glutamate decarboxylase gene and therefore they possibly can be used for industrial lamda-aminobutyric acid production

Messenger RNA expression patterns of neurotrophins during transdifferentiation of stem cells from human-exfoliated deciduous teeth into neural-like cells

Stem cells from Human Exfoliated Deciduous teeth [SHED] have the capability to differentiate into neural cells. Neurotrophins including Nerve Growth Factor [NGF], Brain-Derived Neurotrophic Factor [BDNF], neurotrophin-3 [NT-3], and neurotrophin-4 [NT-4] have neurogenesis, neurotrophic, or neuroprotective effects and are expressed in developing teeth. The aim of this study was to measure quantitative changes in mRNA expression levels of neurotrophins in neural-like cells differentiated from dental pulp stem cells. Isolated total RNA from SHED, dental pulp and neural-like cells [n=3] were transcribed into cDNA. Then real time PCR was done. Expression levels of mRNA for NGF, BDNF, NT-3, and NT-4 genes were compared in these three cells. In neural like cells, BDNF mRNA increased [372.1 +/- 113.5] significantly [p<0.01] after differentiation. NGF mRNA increased to more than 266 times the dental pulp level after differentiation. A similar pattern was seen for the expression of NT3 after differentiation. NT4 mRNA enhancement was 1344 +/- 630.8 and 30.7 +/- 7.9 fold in neural like cells and SHED cells, respectively. Results show alterations with different degrees and direction in neurotrophins mRNA expression levels in these cells. Our results suggest that neurotrophins dental pulp cells, SHED cells and neural like cells derived from SHED cells produce neurotrophic factors. Since the large amounts of neurotrophins are expressed in SHED and neural like cells they may have important role in survival and differentiation of dental pulp stem cells and obtained information may lead to a novel method for tooth regeneration

GABA[A] receptors as novel drug targets for treatment of mental disorders

A balance between excitatory and inhibitory neurotransmissions in brain is an essential factor for the proper function of the brain. The amino acid gamma-aminobutyric-acid [GABA] is considered as the major inhibitory neurotransmitter in brain. Thus, GABAergic neurons play a key role in regulating behavior. Previous data have revealed the complex subunit structural design for GABAA receptor channel, in which a pentameric assembly resulting from 5 of at least 21 subunits, grouped in the eight classes alpha [[alpha]1-6], beta [[beta]1-4], gamma [[gamma]1-4], delta, pi [[PI]], epsilon [[epsilon]], theta [theta] and rho [rho1-3] permits an immense number of putative receptor isoforms. GABAARs are highly diverted in the central nervous system in which this diversity may be related to some mental disorders. Any alteration in expression of the GABAA receptor genes causes neurophysiological and functional consequences that might be associated with neurological disorders. Some neuropsychiatric disorders, such as anxiety, epilepsy and sleep disorders, are effectively treated with therapeutic agents that act on the GABAA receptor. In this article, the contribution of GABAA receptor deficits to central nervous system disorders, in particular anxiety disorders, epilepsy, schizophrenia and insomnia, will be reviewed. The better understanding of GABA and its receptors may help us to find novel therapeutic agents for treatment of mental disorder in future research

Effects of rat's licking behavior on cutaneous wound healing

Wound licking has been shown to advance wound healing among humans and many other animals. The present study evaluates the licking effects on healing of skin wound in rats. Twenty four rats were assigned to 4 different groups randomly and two 3 cm longitudinal full thickness incisions were made on each dorsal and ventral side of rats. The ventral incisions were considered as treated wounds because of contact to saliva as rats lick them easily and dorsal incisions as control wounds. Clinical changes and histopathological effects of rat saliva on wound healing were evaluated every day and on 3, 7, 14 and 21 days post-operation respectively. Histologic and clinical evaluation of treated wounds showed better healing than control wounds. This study showed that licking behavior can promote wound healing. Thus salivary compounds could be isolated, be mass produced and may have potential to become as common as antibiotic cream

[Application of adeno-associated virus [AAV] as a gene therapy vector]

Adeno-associated virus [AAV] is a small virus encapsidates a single-stranded DNA genome. AAV have rapidly gained popularity as a vector in genetic engineering and gene therapy applications, due to its lack of pathogencity, wide range of infectivity, and ability to integrate into human genome [19q13.4]. Recombinant AAV vectors have been used for treatment of a variety of diseases such as hemophilia, cystic fibrosis, and Parkinson. A challenge with wide application of this vector is the relatively small packaging size of the viral genome to enable the delivery of a large transgene. Therefore it is essential making changes in it to overcome this problem. In this review we will introduce with this virus and the new finding in reginal of gene therapy with this virus