Adenosine A_2A Receptor Agonist Polydeoxyribonucleotide Alleviates Interstitial Cystitis-Induced Voiding Dysfunction by Suppressing Inflammation and Apoptosis in Rats

Figures & data

Figure 1 Experimental schedule.

Table 1 Histological Score

Score	Histological Characteristics
0	Morphologically unremarkable with no or very minimal inflammation or epithelial changes.
1	Minimal inflammatory infiltrate composed of occasional neutrophils or lymphocytes within the lamina propria in the absence of inflammation in the muscularis propria, or significant edema, hemorrhage or urothelial changes.
2	Minimal to mild inflammatory infiltrate within the lamina propria with scattered neutrophils or lymphocytes, accompanied by mild edema or hemorrhage, but in the absence of inflammation in the muscularis propria or significant urothelial changes.
3	Mild or mild to moderate inflammatory infiltrate in the lamina propria and focal extension of the inflammation into the muscularis propria.
4	Moderate inflammation with scattered to frequent neutrophils and lymphocytes in both the lamina propria and muscularis propria.
5	Severe inflammation in the lamina propria and muscularis propria in association with other significant findings, such as urothelial ulceration, severe edema, hemorrhage and fibrin deposition.

Table 2 Primary and Secondary Antibodies in Western Blot

Classification	Items	Source	Titer	Company
Primary antibody	NF-κB	Anti-rabbit	1:1000	Abcam, Cambridge, UK
	IκB-α, p-IκB-α, CREB, p-CREB, PKA, p-PKA	Anti-rabbit	1:1000	Santa Cruz Biotechnology, CA, USA
	ERK, p-ERK, JNK, p-JNK, p38, p-p38	Anti-rabbit	1:2000	Cell Signaling Technology, Danvers, USA
	Bax, Bcl-2, β-actin	Anti-mouse	1:1000	Santa Cruz Biotechnology, CA, USA
Secondary antibody	HRP-conjugated IgG	Mouse	1:2000	Vector Laboratories, CA, USA
		Rabbit

Abbreviation: p, phosphorylation.

Figure 2 Voiding functions. (A) The inter-contraction pressure, contraction pressure and time change as recorded by cystometry. (B) Comparison of inter-contraction interval in each group. (C) Comparison of contraction pressure in each group, (D) Comparison of contraction time in each group. *Indicates P < 0.05 when compared with the control group. ^#Indicates P < 0.05 compared with the IC-induced group.

Figure 3 Ratio of bladder weight to body weight and inflammatory score. (A) Histological changes in the bladder tissue. (B) Ratio of bladder weight to body weight in each group. (C) Inflammatory score in each group. Black arrows indicate epithelial detachment, erosion, and inflammatory cells. Blue arrows indicate edematous stroma and irregularly ectasia vessels. *Indicates P < 0.05 when compared with the control group. ^#Indicates P < 0.05 compared with the IC-induced group.

Figure 4 Pro-inflammatory cytokines. Left: Tumor necrosis factor (TNF)-α level in the bladder tissue. Right: Interleukin (IL)-1β level in the bladder tissue. *Indicates P < 0.05 when compared with the control group. ^#Indicates P < 0.05 compared with the IC-induced group.

Figure 5 Nnuclear factor (NF)-κB, NF-κB inhibitor-α (IκB-α), and cascade factors of mitogen-activated protein kinase (MAPK). Upper left panel: Representative expressions of NF-κB, NF-κB inhibitor (IκB)-α, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p-38. Upper right panel: Relative expression of NF-κB and relative ratio of phosphorylated IκB-α (p-IκB-α) to IκB-α. Lower panel: Relative ratio of phosphorylated ERK (p-ERK) to ERK, phosphorylated JNK (p-JNK) to JNK, and phosphorylated p-38 (p-p-38) to p-38. *Indicates P < 0.05 when compared with the control group. ^#Indicates P < 0.05 compared with the IC-induced group.

Figure 6 Cyclic adenosine-3ʹ,5ʹ-monophosphate (cAMP), cAMP response element-binding protein (CREB) and protein kinase A (PKA). (A) cAMP level in the bladder tissue. (B) Relative ratio of phosphorylated CREP (p-CREB) to CREB in bladder tissue. (C) Relative ratio of phosphorylated PKA (p-PKA) to PKA in bladder tissue. *Indicates P < 0.05 when compared with the control group. ^#Indicates P < 0.05 compared with the IC-induced group.

Figure 7 Apoptosis in the bladder tissue. (A) Percentages of terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL)-positive (red arrows) cells in the bladder tissue. (B) Relative ratio of Bax to Bcl-2 in bladder tissue. *Indicates P < 0.05 compared with the control group. ^#Indicates P < 0.05 compared with the IC-induced group.