Ruboxistaurin hydrochloride (LY 333531 hydrochloride) is an Isozyme-selective inhibitor of protein kinase C (PKC); competitively and reversibly inhibits PKCβI and PKCβII (IC50 values are 4.7 and 5.9 nM respectively). Selective for PKCβ over other PKC isozymes (IC50values are 0.052, 0.25, 0.30, 0.36, 0.60 and >100 μM for PKCη, -δ, -γ, -α, -ε and -ζ respectively). Exhibits selectivity for PKC over other ATP-dependent kinases, including protein kinase A, casein kinase and src).

PKCη:0.052 μM, PKCβ2:5.9 nM, PKCδ:0.25 μM, PKCβ1:4.7 nM, PKCγ:0.3 μM

Ruboxistaurin显著降低HUVEC存活率，并且与AGEs组相比，Ruboxistaurin在促进HUVEC细胞凋亡方面的效果显著增强。PKC-beta的阻断上调Bax和Bad蛋白表达，下调Bcl-2蛋白表达。此外，Ruboxistaurin降低Bcl-2/Bax比率。Ruboxistaurin还能进一步促进AGEs诱导的内皮细胞凋亡。Bax、Bad的表达增加和Bcl-2的表达减少以及Bcl-2/Bax比率降低与凋亡过程相关[3]。

在糖尿病大鼠中观察到TGF-b1、Smad2和Smad3 mRNA表达的显著上调，而通过给予ruboxistaurin治疗后，这一现象得到了缓解。

HUVECs are seeded into 96-well plates in low glucose DMEM with 10% FBS for 12 h. Afterwards, HUVECs are starved for 12 h and incubated with BSA (200 μg/ml), AGEs (200 μg/ml) and LY333531 (200 nM)+AGEs (200 μg/ml) for 48 h. Then, the medium is replaced with 0.5 mg/ml MTT and at 37 ?C in a 95% air/5% CO2 incubator for 4 h. Finally, the medium containing MTT is aspirated and replaced by dimethyl sulphoxide (DMSO). OD is measured with a Microplate spectrophotometer. AGEs:advanced glycation end products.(Only for Reference)