High mobility group box 1 contributes to wound healing induced by inhibition of dipeptidylpeptidase 4 in cultured keratinocytes.

Dipeptidyl peptidase 4 (DPP4) is expressed in various tissues, including the skin, and DPP4 inhibitors, that are currently used for the treatment of diabetes, may be effective also for complications of diabetes that affect the skin. To assess the role of DPP4 in keratinocytes, after creating a scratch wound in a monolayer of NTCC 2544 cells, we evaluated DPP4 expression and monitored wound repair over time, after treatment with the DPP4 inhibitor 1(((1-(hydroxymethyl)cyclopentyl)amino)acetyl)2,5-cis-pyrrolidinedicarbonitrile (DPP4-In). Expression of DPP4 increased early and was maintained up to 48 h following the scratch as shown by western blot and immunostaining. Treatment with 10 µM DPP4-In reduced DPP4 expression and significantly accelerated wound repair. This effect did not involve enhanced cell proliferation as shown by MTT proliferation assay, the lack of changes of cell cycle profiles and the slight inhibition of ERK phosphorylation. Enhancement of wound repair by DPP4 inhibition was prevented by the non-specific MMPs inhibitor GM6100 (5 µM). Treatment with DPP4-In increased the expression of high mobility group box 1 (HMGB1), a substrate of this enzyme, and exposure of NCTC 2544 cells to DPP4-In and exogenous HMGB1 (10 nM) produced a non-additive effect. Finally the healing promoting effect of DPP4-In was prevented by pretreatment with a neutralizing anti-HMGB1 antibody. The present results suggest that DPP4 inhibition contributes to enhanced wound healing by inducing keratinocytes to migrate into a scratched area. This effect seems to be independent of cell proliferation and involves enhanced production of HMGB1.

Pharmacokinetic characterization of tizanidine nasal spray, a novel intranasal delivery method for the treatment of skeletal muscle spasm.

BACKGROUND AND OBJECTIVE: The skeletal muscle relaxant tizanidine is approved by the US FDA and the European Medicines Agency for treating spasticity and is supplied as tablets for oral administration. However, tizanidine has a poor bioavailability, due to extensive first-pass metabolism. Therefore, the nasal route of administration, which bypasses portal circulation, may increase the bioavailability of tizanidine and, possibly, reduce the time to peak plasma concentration, thereby shorting the latency of therapeutic effect. The objective of this study was to evaluate the pharmacokinetic profile of tizanidine nasal spray and compare it to the profile of tizanidine oral tablets. METHODS: This open-label, phase I study comprised two protocols: protocol 1, tizanidine HCl solution (32.73 mg/mL) intranasally at single doses of 2 and 4 mg versus 4 mg tizanidine oral tablets (randomized, three periods crossover, 12 healthy subjects); and protocol 2, tizanidine HCl solution (16.36 mg/mL) intranasally at a single dose of 1 mg vs. 4 mg tizanidine oral tablets (randomized, two periods crossover, 12 healthy subjects, one dropout). Tizanidine plasma concentrations were determined by liquid chromatography/mass spectrometry. RESULTS: There was a linear relationship between different dosages of intranasal formulation and the area under the concentration-time curve and maximum plasma concentration (C(max)). The relative bioavailability of the different dosages of intranasal formulation were 1.29, 1.93, and 4.23 for 1, 2, and 4 mg intranasal administration, respectively. Comparison of C(max) values gave the following ratios: 0.91, 1.39, and 2.73, for 1, 2, and 4 mg intranasal administration, respectively. The mean time to C(max) (t(max)) was 0.99, 0.43, and 0.63 h for 1, 2, and 4 mg intranasal administration, respectively, whereas it was 1.13 and 1.30 h for the two series of 4 mg tizanidine oral tablets. CONCLUSIONS: The bioavailability of the tizanidine intranasal formulation was higher than that of tizanidine oral tablets. The t(max) was also shorter with the intranasal formulation. No serious adverse events occurred throughout the study, such that the two formulations resulted equally well-tolerated. The intranasal formulation of tizanidine results are therefore worthy of subsequent clinical testing in phase II.

Linagliptin: A thorough Characterization beyond Its Clinical Efficacy.

Linagliptin, one of the five dipeptidyl peptidase-4 inhibitors available, has recently entered the market both in the US and in most European countries for treatment of type 2 diabetes mellitus. It presents a xanthine-based structure, and is characterized by unique pharmacokinetics, with non-linear profile, long terminal half-life allowing prolonged exposure to the drug. It is eliminated predominately through the intestinal tract and only minimally into urine, so that it can be administered, without any dose adjustment, in conditions of renal impairment. Linagliptin is effective in modifying all parameters of hyperglycemia either in monotherapy, or as add-on therapy, together with metformin or a sulfonylurea. It also exhibits a good tolerability profile with few side effects, absence (when used in monotherapy), or low risk (when in combination with a sulfonylurea) of hypoglycemia. More importantly it has a weight neutral effect. A comprehensive report of the literature on linagliptin is provided, paying attention in particular to preclinical studies, interactions with other drugs, safety and tolerability, and results obtained in animal models that highlight properties of linagliptin suggestive of potential additional uses. Particularly promising appear the data demonstrating a positive effect of linagliptin on metabolic dysfunction and renal and/or cardiovascular damage together with more recently reported effects of linagliptin on tissue repair and neuroprotection.

Reversible inhibition of vasoconstriction by thiazolidinediones related to PI3K/Akt inhibition in vascular smooth muscle cells.

Thiazolidinediones (also referred to as glitazones), agonists for Peroxisome Proliferator-Activated Receptor gamma (PPARγ), are used for treating type 2 diabetes mellitus, where they decrease insulin resistance and cardiovascular risk. Compounds bearing the thiazolidinedione structure have also been shown to inhibit phosphoinositide 3-kinase (PI3K). Here we tried to elucidate the poorly defined role of PI3K/Akt in the physiology of vascular smooth muscle cell contraction and tested the hypothesis that thiazolidinediones, by affecting the PI3K/Akt pathway, may influence vascular physiology. Isolated rat femoral arteries segments were mounted in a wire myograph and challenged with 100mM KCl or phenylephrine (PE), in the absence or presence of troglitazone, rosiglitazone, pioglitazone, LY294002 (PI3K inhibitor) and 10-DEBC (Akt inhibitor). All these compounds dose-dependently inhibited vasoconstriction to KCl or PE; their effect was reversible (after 60-120 min washout) and not affected by GW9662 (a PPARγ antagonist) or by N(G)-nitro-L-arginine (LNNA, an inhibitor of NO biosynthesis). Analysis of phospho-Akt (ser 473) in lysates from rat arteries (by immunoblot) revealed that thiazolidinediones, LY294002 and 10-DEBC, at the same concentration and kinetics inhibiting vasoconstriction, produced a similar decrease of Akt phosphorylation. PI3K/Akt pathway therefore appears to be an important, fast acting, modulator of contraction of vascular smooth muscle. Thiazolidinediones decrease vasoconstriction of isolated vessels possibly by inhibiting PI3K/Akt pathway. Such an effect of glitazones, if occurring in vivo, may impact cardiovascular syndromes related to vasospasm in diabetic patients.

Endothelium-dependent vasomotor effects of telmisartan in isolated rat femoral arteries.

AT(1) receptor antagonists (ARBs) are drugs widely used for preventing and/or treating major cardiovascular diseases. Some of these drugs also show AT(1) receptor-independent effects that may have patho-physiological significance, such as Peroxisome Proliferator-Activated Receptors gamma (PPARγ) stimulation. Here we investigated the effect of telmisartan (that also stimulates PPARγ) on vasomotor responses of femoral arteries isolated from rat, in comparison to losartan. Femoral artery segments were mounted in a wire myograph and challenged with cumulative concentrations of phenylephrine (PE) and acetylcholine (ACh) after 30-min incubation in the absence or presence of 30 µM telmisartan or 30 µM losartan. Vasomotor responses were not significantly changed by losartan, whereas telmisartan reduced vasoconstriction to PE and increased vasodilatation to ACh. Incubation with 0.1 mM N(G)-nitro-l-arginine abolished relaxation to ACh in untreated controls as well as in losartan-treated preparations, but did not in telmisartan-treated preparations (were 20% relaxation subsisted); this residual relaxing effect was abolished by indomethacin and by endothelium removal. Incubation with 30 µM GW9662 (PPARγ antagonist), 10 µM PD123319 (AT(2) antagonist) or 30 µM A779 (angiotensin(1-7)/Mas antagonist) did not change the effect of telmisartan on vasomotor responses in preparations with intact endothelium. We conclude that telmisartan modifies constriction and dilatation of isolated arteries in an endothelium-dependent manner, involving both nitric oxide and prostanoid production. The present effect of telmisartan, however, does not seem to involve PPARγ, AT(2) or angiotensin(1-7)/Mas.