23/05/24, 02:11 Alterações na expressão de múltiplos transcritos de fatores neurotróficos derivados do cérebro no modelo de ligação seletiva d…

Pesquisa cerebral
Volume 1206, 24 de abril de 2008 , páginas 13-19

Relatório de pesquisa

Alterações na expressão de múltiplos transcritos de fatores

neurotróficos derivados do cérebro no modelo seletivo de
ligação do nervo espinhal e no modelo adjuvante completo
de Freund
Hiroyuki Kobayashi e,Masataka Yokoyama e ,Yoshikazu Matsuoka e,Megumi Omori a,Yoshitaro Itano a,Rhuji Kaku a,
Kiyoshi Morita e,Hiroyuki Ichikawa b

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https://doi.org/10.1016/j.brainres.2007.12.004
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Abstrato

Alterações na expressão do fator neurotrófico derivado do cérebro (BDNF) no gânglio da raiz dorsal (DRG) e na medula
espinhal em alguns modelos de dor. Recentemente, transcritos de BDNF de rato contendo novos exons 5' não traduzidos
foram identificados e caracterizados, e um novo sistema de numeração para exons de BDNF de rato foi introduzido.
Examinamos os perfis de expressão desses novos transcritos de BDNF em DRGs L4/5 bilaterais em um modelo de ligação
seletiva do nervo espinhal (SSNL) para L5 e DRGs L5 bilaterais em um modelo completo de adjuvante de Freund (CFA) de
ratos. L5SSNL aumentou significativamente ( P < 0,05) a expressão do mRNA total do BDNF e dos transcritos do exon I,
IIA, IIB, IIC, III, IV, VI e IXA no L4 DRG ipsilateral. Embora a expressão do mRNA total do BDNF tenha permanecido
inalterada no DRG L5 ipsilateral no modelo L5SSNL, a expressão do transcrito do exon I aumentou significativamente ( P
< 0, 05) e a do transcrito do exon IV diminuiu significativamente ( P < 0, 05). Os perfis de expressão dos exões variantes
no DRG L4 ipsilateral do modelo L5SSNL foram bastante semelhantes aos do DRG ipsilateral do modelo CFA, e o
transcrito do exão I foi o ARNm de BDNF mais comum nestes DRGs. Embora L5SSNL tenha aumentado
significativamente ( P < 0, 05) a expressão do mRNA total do BDNF e dos transcritos do exon IIC e IXA nos DRGs L4/5
contralaterais, o tratamento com CFA não alterou a expressão do mRNA total do BDNF ou transcritos específicos nos
DRGs contralaterais. Estes resultados sugerem que o exão I desempenha um papel importante no aumento da expressão
do BDNF em DRGs ipsilaterais, independentemente da condição.

Introdução

Brain-derived neurotrophic factor (BDNF) is a member of the neurotrophin family and plays important roles in survival,
differentiation and synaptic plasticity of neurons. In adult rats, BDNF mRNA and protein are constitutively expressed by
small neurons in the dorsal root ganglion (DRG) (Wetmore and Olson, 1995, Zhou and Rush, 1996), and BDNF protein is
transported to superficial laminae of the spinal dorsal horn (Cho et al., 1997a). The levels of BDNF expression change in
the DRG and spinal cord in some pain models. In the dorsal horn, activation of trkB receptors by endogenous BDNF
appears to contribute to hyperalgesia associated with peripheral inflammation (Woolf and Salter, 2000), and it has been

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suggested that BDNF contributes to the sensitization of dorsal horn neurons in neuropathic pain model (Fukuoka et al.,
2001). In an inflammatory pain model, expression of BDNF mRNA in small neurons in ipsilateral L4/5 DRG and the level
of BDNF protein in the spinal cord are increased by injection of complete Freund's adjuvant (CFA) into the hindpaw (Cho
et al., 1997a, Cho et al., 1997b). In the L5-selective spinal nerve ligation (SSNL) model of rats, however, the profile of
BDNF protein expression between ipsilateral L4 and L5 DRG is different; BDNF levels are increased in L4 small DRG
neurons and L5 large DRG neurons (Ha et al., 2001). This indicates that there are different mechanisms underlying the
changes in BDNF mRNA and protein between L4 and L5 DRGs.

BDNF gene is transcribed into multiple transcripts and expression of the various transcripts is controlled by multiple
activity-dependent and tissue-specific promoters (Timmusk et al., 1993). Recently, Liu et al. (2006) and Aid et al. (2007)
identified and characterized several rodent BDNF transcripts containing novel 5′ untranslated exons and introduced a
new numbering system for rat BDNF exons. According to Aid et al. (2007), rat BDNF gene contains a common 3′ exon
that encodes the pro-BDNF protein (exon IX) and at least eight 5′ noncoding exons (exons I–VIII). Exon II, which was
previously characterized by Timmusk et al. (1993), was subdivided by Aid et al. (2007) into exons IIA, IIB and IIC. Exons
previously named exons III and IV were renamed exons IV and VI, respectively. The coding exon, which was previously
called exon V, was renamed exon IX. Transcription of the gene yields BDNF transcripts containing one of the eight 5′
exons or the 5′ extended coding exon (exon IXA), which is spliced to the coding exon. In total, there are 11 different
noncoding exons that can be spliced to the coding exon to form the following splice variants: exons I–IX (I), IIA–IX (IIA),
IIB–IX (IIB), IIC–IX (IIC), III–IX (III), IV–IX (IV), V–IX (V), VI–IX (VI), VII–IX (VII), VIII–IX (VIII), and IXA–IX (IXA). Exon IX
represented the total BDNF mRNA in the current study.

Kim et al. (2001) examined expression profiles for multiple BDNF transcripts in DRGs in rats subjected to several pain
models. They reported that expression of transcript exons I, II and III (old nomenclature) was increased in inflammatory
pain models. However, the expression profiles of these various BDNF transcripts have not been reported in the L5SSNL
model of rats. Given the suggestion for segmental heterogeneity, it is important to investigate the expression profiles of
BDNF transcripts between L4 and L5 DRGs as well.

Fukuoka et al. (2001) reported that total BDNF mRNA expression in the ipsilateral L4 DRG began to increase significantly
at 3 days after L5SSNL and maintained until 21 days and that BDNF protein also increased significantly at 14 days after
L5SSNL. In addition, the possibility that postoperative inflammation has an effect on BDNF mRNA in the DRG by 7 days
after L5SSN cannot be neglected. In the present study, therefore, we examined expression of the BDNF transcripts using
the new nomenclature in ipsilateral and contralateral L4/5 DRGs at 14 days after in the L5SSNL rat as a neuropathic pain
model. We also examined expression of BDNF transcripts in the CFA rat as an inflammation model for comparison
between the two frequently utilized pain models.

Section snippets

Behavioral assessments of L5SSNL and CFA rats

Behavioral assessments were determined by von Frey test before and after L5SSNL and CFA treatment. Mechanical
allodynia was observed in L5SSNL model rats throughout the examined periods (days 1–14 after L5 spinal nerve
ligation) (Fig. 1). CFA injection induced allodynia at 2 days after the treatment (Fig. 2).…

Expression profiles of multiple BDNF transcripts in intact rats

In intact animals, there was no significant difference in the expression of exon IX (total BDNF mRNA) or any BDNF
transcripts between right and left L4/5 DRGs (Fig. 3, Fig. 4). Exon I, IIA, …

Discussion

The present study demonstrated that L5SSNL changes the expression profiles of BDNF transcripts in L4/5 DRGs, and that
the expression profiles of BDNF transcripts are different between L4 and L5 DRGs. The study also demonstrated that the

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expression profile of the effected exons in ipsilateral L4 DRG of the L5SSNL model is quite similar to those in ipsilateral
DRG of the CFA model. In both models, the exon I transcript is the most common BDNF mRNA in these DRGs.

We found that L5SSNL increased the…

Experimental procedures

All experiments were carried out under the control of the Animal Research Control Committee in accordance with the
Guidelines for Animal Experiments of Okayama University Medical School, Government Animal Protection and
Management Law (No. 105) and the Japanese Government Notification on Feeding and Safekeeping of Animals (No. 6).
All efforts were made to minimize the number of animals used and their suffering.…

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…Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family of proteins, is widely distributed in the pain-related
pathway and limbic system of the central nervous system, suggesting that it is likely to play a crucial role in pain-related functions
(Phillips et al., 1990; McAllister et al., 1999). Numerous studies have reported that peripheral nerve injury upregulates BDNF
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…No presente estudo, experimentos de western blotting mostraram que os níveis de expressão proteica do BDNF maduro foram
regulados positivamente nos gânglios da raiz dorsal e na medula espinhal após lesão nervosa poupada. Nossos dados são consistentes
com estudos anteriores que relataram que a lesão do nervo periférico regula positivamente os níveis de mRNA e proteína do BDNF no
DRG (Fukuoka et al., 2001; Obata et al., 2003; Kobayashi et al., 2008), bem como o BDNF. níveis de proteína no corno dorsal da coluna
vertebral (Miletic e Miletic, 2002; Geng et al., 2010). Após lesão nervosa periférica, o BDNF mostra um papel bem definido na
sobrevivência e expressão fenotípica de neurônios sensoriais primários nos gânglios da raiz dorsal (Anand, 1996).…

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A entrega do gene BDNF mediada por nanopartículas direcionadas a neurônios é neuroprotetora na lesão
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Regulação epigenética da expressão do BDNF nos neurônios sensoriais primários após lesão nervosa
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A regulação da expressão de mRNA do fator neurotrófico derivado do cérebro específico do exon pela
proteína quinase C em neurônios ganglionares da raiz dorsal cultivados em ratos
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…Além disso, a estimulação com ATP leva à regulação positiva do éxon IV, mas não do éxon I, II ou VI, através da quinase dependente
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expressão dos exons I – IV e VI foi demonstrada em neurônios DRG de ratos (Kobayashi et al., 2008). Um estudo recente relata que o
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Estratégia chamariz visando o éxon I do fator neurotrófico derivado do cérebro para atenuar a alodinia tátil
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…Os resultados apresentaram a primeira evidência de que o pós-tratamento intratecal com ODNs chamariz para o promotor do exon I
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acordo com nossos dados anteriores, a expressão do mRNA do exon I de Bdnf aumentou notavelmente nos DRGs L4 e L5 ipsilaterais
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