NPPB (Synonyms: HOE 144, Hoechst 144)

Cell lines

Primary adhesion fibroblasts (AFB)

Preparation Method

Primary peritoneal AFB cells were seeded in 12-well plates at a density of 1.9×10⁵/well and allowed to grow from 80 to 90% confluence. The confluent monolayers were scratched with a 10μL pipette tip and then washed with PBS several times to remove cell debris. The culture medium was replaced with medium containing 1% BSA, 1% FBS, and increasing concentrations of NPPB (0, 30, 60, 90, and 120μM). Wound-healing was photographed using an inverted fluorescence microscope to assess the rate of closure. The migration rates were calculated using the equation [(wound area at 0h−wound area at 72h)/wound area at 0h]×100%. The experimental data at 72h were transformed to sigmoidal dose-response curves using non-linear regression analysis to calculate the corresponding IC₅₀ values.

Reaction Conditions

30–120μM; 72h

Applications

NPPB inhibited the migration of adhesion fibroblasts (AFB) with an IC₅₀ of 56.75μM.

Animal models

Albino rabbits

Preparation Method

Thirty Albino rabbits of both sexes weighting 2.7-3.6kg were divided into five groups. All the animals were anesthetized by 30mg/kg sodium pentobarbital (iv) and ventilated using a respirator. A 5 French high flow multipurpose catheter was placed in the vertebral artery through the femoral artery in each animal. A control angiogram of the basilar artery was performed in each group. Fifteen minutes after the control angiogram, 0.6ml of isotonic saline in Group I, 0.6ml of 1nM ET-1 in Group II were slowly injected into the vertebral arteries of the animals. Thirty minutes after the injection, control angiograms were performed to evaluate the effects of ET-1 and isotonic saline. In the ET-1 plus NPPB treated group (Group III), 0.1ml 20mM NPPB was slowly injected after the control angiogram. 0.6ml, 1nM ET-1 was slowly injected ten minutes after the NPPB administration. The last angiograms were performed thirty minutes after the injection of ET-1 to evaluate the effects of ET-1 in NPPB treated rabbits. In Group IV, the effect of dimethylsulfate (DMS) was tested by a control angiogram thirty minutes after the injection of 0.1ml DMS since NPPB was dissolved in DMS. In Group IV, the effect of NPPB in the normal basilar artery was tested by a control angiogram 30 minutes after the injection of 0.1ml 20mM NPPB. On the subtraction images, the diameter of the basilar artery in pre and post injection angiograms was measured in each group and compared by quantitative analysis software. All animals were sacrificed immediately after the repeat angiography by exsanguination. Basilar arteries were rapidly removed and processed by methods described previously for ultrastructural investigation.

Dosage form

0.1ml; 20mM; intra-arterial injection

Applications

NPPB prevented ET-1-induced vasoconstriction and reversed ET-1-induced histological alterations in the brainstem of rabbits.

References:
[1] Zhong J, Qin Z, Yu H, et al. NPPB prevents postoperative peritoneal adhesion formation by blocking volume-activated Cl^- current. Naunyn Schmiedebergs Arch Pharmacol. 2020;393(3):501-510.
[2] Dogulu F, Barun S, Emmez H, et al. Effect of a chloride channel inhibitor, 5-nitro-2- (3-phenylpropylamino)-benzoate, on endothelin-1 induced vasoconstriction in rabbit basilar artery. Turk Neurosurg. 2009;19(4):380-386.