Herpes simplex virus mediated nerve growth factor expression in bladder and afferent neurons: potential treatment for diabetic bladder dysfunction.
OBJECTIVE
Diabetic cystopathy resulting from sensory neuropathy may potentially be treated by direct gene therapy. It has been suggested that nerve growth factor (NGF) has an ameliorative effect in preventing the death in diabetes of afferent dorsal root ganglion neurons, which control bladder function. We investigated NGF gene transfer to the bladder and bladder afferent pathways for treating diabetic cystopathy. We used replication competent and replication defective herpes simplex virus type 1 (HSV-1) vectors that express a functionally active form of the beta-subunit of mouse NGF (beta-NGF) to examine the level and duration of therapeutic gene expression after administration of the vectors.


METHODS
NGF expression during acute (3 days) and latent (21 days) infections was assessed by enzyme-linked immunosorbent assay (ELISA) and immunohistochemical testing after the injection of 1 x 106 to 1 x 108 pfu HSV-NGF expression vectors into the bladder wall of adult rats.


RESULTS
HSV vectors with the strong human cytomegalovirus immediate early promoter used to drive beta-NGF gene expression exhibited increased NGF 3 days after infection in the bladder and L6 to S1 dorsal root ganglia, where bladder afferent neurons are located. ELISA analysis revealed that NGF in the bladder tissue and dorsal root ganglia was increased 7 to 9 and 2 to 4-fold, respectively, over the control vector. Increased NGF expression in L6 to S1 dorsal root ganglia neurons was also detected by immunohistochemical staining with antiNGF antibodies. Extended NGF expression was detected by ELISA 21 days after injection. Replication defective vectors containing HSV-1 latency promoter (LAP-2) driving NGF expressed NGF in the bladder and dorsal root ganglia 21 days after bladder injection. ELISA analysis confirmed an approximate 2 to 3-fold increase of NGF expression in the bladder and L6 to S1 dorsal root ganglia.


CONCLUSIONS
The NGF gene may be transferred and expressed in the bladder and bladder afferent pathways using HSV vectors. To our knowledge our study represents the first demonstration of the effectiveness of gene therapy for altering neurotrophic expression in visceral sensory neurons. This technique of gene transfer may be useful for treating certain types of neurogenic bladder dysfunction, such as diabetic cystopathy, in which decreased NGF transport may be a causative factor.
Goins
W F
WF
Department of Urology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA.
Yoshimura
N
N
Phelan
M W
MW
Yokoyama
T
T
Fraser
M O
MO
Ozawa
H
H
Bennett N
J R
JR
de Groat
W C
WC
Glorioso
J C
JC
Chancellor
M B
MB
D000818:Animals; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism*; D003929:Diabetic Neuropathies / Q000150:complications*; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005455:Fluorescent Antibody Technique; D005727:Ganglia, Spinal / Q000378:metabolism; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D005822:Genetic Vectors; D018259:Herpesvirus 1, Human / Q000235:genetics*; D020932:Nerve Growth Factor / Q000235:genetics / Q000378:metabolism*; D009475:Neurons, Afferent / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D001743:Urinary Bladder / Q000294:innervation / Q000378:metabolism*; D001750:Urinary Bladder, Neurogenic / Q000209:etiology / Q000378:metabolism / Q000628:therapy*
D020932:Nerve Growth Factor