The aim of this study was to evaluate whether
liposome-based local suppression of
nerve growth factor (
NGF) in the bladder has effects on bladder
hypersensitivity in a rat
cystitis model induced by
intravesical instillation of
hydrogen peroxide (HP). HP (1.5%) was intravesically administered to adult female Sprague-Dawley rats.
Liposomes complexed with
NGF antisense oligonucleotide (OND) labeled with TYE563 fluorescent tag were intravesically instilled on day 2. Red fluorescence from the TYE 563 tag was observed with fluorescent microscopy on day 3. Four separate groups of rats were used in the following experiments: (a)
sham-
liposome group, (b)
sham-OND group, (c)
cystitis-
liposome group, and (d)
cystitis-OND group. Saline or 1.5% HP was intravesically administered on day 0. Empty
liposomes or
liposomes-antisense OND were instilled into the bladder on day 2. The following experiments were conducted to evaluate the effect of
NGF antisense treatment on day 7: (a) continuous cystometry was performed in an awake condition; (b)
pain behavior induced by instillation of
resiniferatoxin into the bladder, including licking behavior (lower abdominal licking) and freezing behavior (motionless head-turning toward lower abdomen), was observed; (c) immunohistochemical staining of the bladder and L6 DRG for
NGF was performed; (d) the expression of several genes in the bladder was analyzed by reverse transcription polymerase chain reaction (RT-PCR); and (e) after
Fast Blue was injected into the bladder wall,
Fast Blue-positive or -negative cells in DRG neurons were separately collected by using a
laser-capture microdissection method 7 days later. RT-PCR was performed to evaluate gene expressions in captured neuronal cells. The expression of TYE563 was identified only in the urothelial layer. In cystometric investigation, intercontraction intervals (ICI) were significantly (p = 0.001) shorter in the
cystitis-
liposome group in comparison to the
sham-
liposome group. ICI was significantly (p = 0.007) longer in the
cystitis-OND group compared to the
cystitis-
liposome group. Comparisons of the
sham-
liposome and the
sham-OND groups showed no significant difference in ICI (p = 0.56). Licking events did not significantly differ among the four groups. In contrast, the
cystitis-
liposome group showed significantly more freezing events than the
sham-
liposome group did (p = 0.002). A significant reduction in the number of freezing events was observed in the
cystitis-OND group compared to the
cystitis-
liposome group (p = 0.04). Immunofluorescence staining demonstrated that
NGF expression in the mucosa (p = 0.02) and L6 DRG (p = 0.01) was significantly higher in the
cystitis-
liposome group than it was in the
sham-
liposome group. The expression of
NGF was significantly lower in the mucosa (p = 0.002) and L6 DRG (p = 0.01) in the
cystitis-OND group compared to the
cystitis-
liposome group. RT-PCR showed that the expression of
NGF and TRPV1
mRNA in the mucosa was significantly higher in the
cystitis-
liposome group than it was in the
sham-
liposome group (p = 0.001 and 0.03, respectively). On the other hand, these gene expressions were significantly lower in the
cystitis-OND group than they were in the
cystitis-
liposome group (p = 0.007 and 0.02, respectively). The
cystitis-
liposome group showed significantly higher expression of TRPA1, P2X3, and
BDNF mRNA in labeled bladder afferent neurons than the
sham-
liposome group did (p = 0.03, 0.01, and 0.001, respectively). These gene expressions were significantly lower in the
cystitis-OND group compared to the
cystitis-
liposome group (p = 0.04, 0.006, and 0.03, respectively). The study indicated that intravesical application of
liposome-
NGF antisense OND significantly improved bladder
hypersensitivity induced by chemical
cystitis in rats. Intravesical treatment with
liposome-OND conjugates could be a novel local
therapy of hypersensitive bladder disorders such as
bladder pain syndrome/
interstitial cystitis.