scholarly journals Advanced Microparticulate/Nanoparticulate Respirable Dry Powders of a Selective RhoA/Rho Kinase (Rock) Inhibitor for Targeted Pulmonary Inhalation Aerosol Delivery

Pharmaceutics ◽  
2021 ◽  
Vol 13 (12) ◽  
pp. 2188
Author(s):  
Priya Muralidharan ◽  
Don Hayes ◽  
Jeffrey R. Fineman ◽  
Stephen M. Black ◽  
Heidi M. Mansour
Keyword(s):  
Pulmonary Hypertension ◽  
Physicochemical Characterization ◽  
Rho Kinase ◽  
Drug Dose ◽  
Particle Engineering ◽  
Rock Inhibitor ◽  
Medical Needs ◽  
Human Lung Cells ◽  
Aerosol Dispersion

Pulmonary hypertension (PH) is a progressive disease that eventually leads to heart failure and potentially death for some patients. There are many unique advantages to treating pulmonary diseases directly and non-invasively by inhalation aerosols and dry powder inhalers (DPIs) possess additional unique advantages. There continues to be significant unmet medical needs in the effective treatment of PH that target the underlying mechanisms. To date, there is no FDA-approved DPI indicated for the treatment of PH. Fasudil is a novel RhoA/Rho kinase (ROCK) inhibitor that has shown great potential in effectively treating pulmonary hypertension. This systematic study is the first to report on the design and development of DPI formulations comprised of respirable nanoparticles/microparticles using particle engineering design by advanced spray drying. In addition, comprehensive physicochemical characterization, in vitro aerosol aerosol dispersion performance with different types of human DPI devices, in vitro cell-drug dose response cell viability of different human respiratory cells from distinct lung regions, and in vitro transepithelial electrical resistance (TEER) as air-interface culture (AIC) demonstrated that these innovative DPI fasudil formulations are safe on human lung cells and have high aerosol dispersion performance properties.

scholarly journals Advanced therapeutic inhalation aerosols of a Nrf2 activator and RhoA/Rho kinase (ROCK) inhibitor for targeted pulmonary drug delivery in pulmonary hypertension: design, characterization, aerosolization, in vitro 2D/3D human lung cell cultures, and in vivo efficacy

2021 ◽  
Vol 15 ◽  
pp. 175346662199824
Author(s):  
Maria F. Acosta ◽  
Priya Muralidharan ◽  
Carissa L. Grijalva ◽  
Michael D. Abrahamson ◽  
Don Hayes ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Engineering Design ◽  
Rho Kinase ◽  
Residual Water ◽  
Particle Engineering ◽  
Rock Inhibitor ◽  
Molecular Fingerprint ◽  
Nrf2 Activator

Inhalable nanostructured microparticles of simvastatin, a Nrf2 activator and RhoA/Rho kinase (ROCK) inhibitor, were rationally designed for targeted pulmonary delivery as dry powder inhalers (DPIs) for the treatment of pulmonary hypertension (PH). Advanced particle engineering design technology was employed to develop inhalable dry powders using different dilute feed concentrations and spray drying pump rates. Several analytical techniques were used comprehensively to characterize the physicochemical properties of the resulting powders. Scanning electron microscopy (SEM) was used to visualize particle morphology (shape), surface structure, size, and size distribution. Karl Fischer titration (KFT) was employed to quantify the residual water content in the powders. X-ray powder diffraction (XRPD) was used to determine crystallinity. Hot-stage microscopy (HSM) under cross-polarizing lens was used to observe the presence or absence of birefringence characteristic of crystallinity. Differential scanning calorimetry (DSC) was employed to quantify thermotropic phase behavior. Attenuated total reflectance (ATR)-Fourier-transform infrared (FTIR) spectroscopy and Raman spectroscopy were used to determine the molecular fingerprint of simvastatin powders before and after particle engineering design. In vitro aerosol dispersion performance was performed with three different Food and Drug Administration (FDA)-approved human DPI devices. Cell viability and transepithelial electrical resistance (TEER) were demonstrated using different in vitro human pulmonary cell two and three-dimensional models at the air–liquid interface, and in vivo safety in healthy rats by inhalation. Efficacy was demonstrated in the in vivo lamb model of PH. Four different inhalable powders of simvastatin were successfully produced. They possessed nanostructured surfaces and were in the inhalable size range. Simvastatin retained its crystallinity following particle engineering design. The more dilute feed concentration spray dried at the lower pump rate produced the smallest particles. All powders successfully aerosolized with all three DPI human devices. Inhaled simvastatin as an aerosol restored the endothelial function in the shunt lamb model of PH, as demonstrated by the reduction of pulmonary vascular resistance (PVR) in response to the endothelium-dependent vasodilator acetylcholine. The reviews of this paper are available via the supplemental material section.

scholarly journals Endothelin-1 impairs angiogenesis in vitro through Rho-kinase activation after chronic intrauterine pulmonary hypertension in fetal sheep

2012 ◽  
Vol 73 (3) ◽  
pp. 252-262 ◽  
Author(s):  
Jason Gien ◽  
Nancy Tseng ◽  
Gregory Seedorf ◽  
Gates Roe ◽  
Steven H. Abman
Keyword(s):  
Pulmonary Hypertension ◽  
Rho Kinase ◽  
Fetal Sheep ◽  
Kinase Activation ◽  
Endothelin 1

scholarly journals Effects of Short-Term Inhibition of Rho Kinase on Dromedary Camel Oocyte In Vitro Maturation

Animals ◽  
2020 ◽  
Vol 10 (5) ◽  
pp. 750
Author(s):  
Hammed A. Tukur ◽  
Riyadh S. Aljumaah ◽  
Ayman Abdel-Aziz Swelum ◽  
Abdullah N. Alowaimer ◽  
Mutassim Abdelrahman ◽  
...  
Keyword(s):  
In Vitro Maturation ◽  
Rho Kinase ◽  
Positive Outcome ◽  
Dromedary Camel ◽  
Second Phase ◽  
Short Term ◽  
Rock Inhibitor ◽  
Nuclear Maturation ◽  
Significant Difference

This is the first report on a biphasic in vitro maturation (IVM) approach with a meiotic inhibitor to improve dromedary camel IVM. Spontaneous meiotic resumption poses a major setback for in vitro matured oocytes. The overall objective of this study was to improve in vitro maturation of dromedary camel oocytes using ROCK inhibitor (Y-27632) in a biphasic IVM to prevent spontaneous meiotic resumption. In the first experiment, we cultured immature cumulus–oocyte complexes (COCs, n = 375) in a prematuration medium supplemented with ROCK inhibitor (RI) for 2 h, 4 h, 6 h, and 24 h before submission to normal in vitro maturation to complete 28 h. The control was cultured for 28 h in the absence of RI. In the first phase of experiment two, we cultured COCs (n = 480) in the presence or absence (control) of RI for 2 h, 4 h, 6 h, and 24 h, and conducted real-time relative quantitative PCR (qPCR) on selected mRNA transcripts. The same was done in the second phase, but qPCR was done after completion of normal IVM. Assessment of nuclear maturation showed that pre-IVM for 4 h yielded an increase in MII oocyte (54.67% vs. 26.6% of control; p < 0.05). As expected, the same group showed the highest degree (2) of cumulus expansion. In experiment 2, qPCR results showed significantly higher expression of ACTB and BCL2 in the RI group treated for 4 h when compared with the other groups. However, their relative quantification after biphasic IVM did not reveal any significant difference, except for the positive response of BCL2 and BAX/BCL2 ratio after 4 and 6 h biphasic IVM. In conclusion, RI prevents premature oocyte maturation and gave a significantly positive outcome during the 4 h treatment. This finding is a paradigm for future investigation on dromedary camel biphasic IVM and for improving the outcome of IVM in this species.

scholarly journals The Vasopressin Receptor 2 Mutant R137L Linked to the Nephrogenic Syndrome of Inappropriate Antidiuresis (NSIAD) Signals through an Alternative Pathway that Increases AQP2 Membrane Targeting Independently of S256 Phosphorylation

Cells ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 1354
Author(s):  
Marianna Ranieri ◽  
Maria Venneri ◽  
Tommaso Pellegrino ◽  
Mariangela Centrone ◽  
Annarita Di Mise ◽  
...  
Keyword(s):  
Water Permeability ◽  
Collecting Duct ◽  
Alternative Pathway ◽  
Water Channel ◽  
Rho Kinase ◽  
Membrane Targeting ◽  
Rock Inhibitor ◽  
Activating Mutations ◽  
Osmotic Water

NSIAD is a rare X-linked condition, caused by activating mutations in the AVPR2 gene coding for the vasopressin V2 receptor (V2R) associated with hyponatremia, despite undetectable plasma vasopressin levels. We have recently provided in vitro evidence that, compared to V2R-wt, expression of activating V2R mutations R137L, R137C and F229V cause a constitutive redistribution of the AQP2 water channel to the plasma membrane, higher basal water permeability and significantly higher basal levels of p256-AQP2 in the F229V mutant but not in R137L or R137C. In this study, V2R mutations were expressed in collecting duct principal cells and the associated signalling was dissected. V2R-R137L and R137C mutants had significantly higher basal pT269-AQP2 levels -independently of S256 and PKA-which were reduced to control by treatment with Rho kinase (ROCK) inhibitor. Interestingly, ROCK activity was found significantly higher in V2R-R137L along with activation of the Gα12/13–Rho–ROCK pathway. Of note, inhibition of ROCK reduced the basal elevated osmotic water permeability to control. To conclude, our data demonstrate for the first time that the gain-of-function mutation of the V2R, R137L causing NSIAD, signals through an alternative PKA-independent pathway that increases AQP2 membrane targeting through ROCK-induced phosphorylation at S/T269 independently of S256 of AQP2.

Acute vasodilator effects of Rho-kinase inhibitors in neonatal rats with pulmonary hypertension unresponsive to nitric oxide

2008 ◽  
Vol 294 (2) ◽  
pp. L205-L213 ◽  
Author(s):  
Patrick J. McNamara ◽  
Prashanth Murthy ◽  
Crystal Kantores ◽  
Lilian Teixeira ◽  
Doreen Engelberts ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Pulmonary Hypertension ◽  
Kinase Inhibitors ◽  
Pulmonary Arteries ◽  
Rho Kinase ◽  
Neonatal Rat ◽  
Rock Inhibitor ◽  
No Donor ◽  
Rat Models ◽  
Pulmonary Hemodynamic

Pulmonary hypertension (PHT) in neonates is often refractory to the current best therapy, inhaled nitric oxide (NO). The utility of a new class of pulmonary vasodilators, Rho-kinase (ROCK) inhibitors, has not been examined in neonatal animals. Our objective was to examine the activity and expression of RhoA/ROCK in normal and injured pulmonary arteries and to determine the short-term pulmonary hemodynamic (assessed by pulse wave Doppler) effects of ROCK inhibitors (15 mg/kg ip Y-27632 or 30 mg/kg ip fasudil) in two neonatal rat models of chronic PHT with pulmonary vascular remodeling (chronic hypoxia, 0.13 FiO2, or 1 mg·kg−1·day−1 ip chronic bleomycin for 14 days from birth). Activity of the RhoA/ROCK pathway and ROCK expression were increased in hypoxia- and bleomycin-induced PHT. In both models, severe PHT [characterized by raised pulmonary vascular resistance (PVR) and impaired right ventricular (RV) performance] did not respond acutely to inhaled NO (20 ppm for 15 min) or to a single bolus of a NO donor, 3-morpholinosydnonimine hydrochloride (SIN-1; 2 μg/kg ip). In contrast, a single intraperitoneal bolus of either ROCK inhibitor (Y-27632 or fasudil) completely normalized PVR but had no acute effect on RV performance. ROCK-mediated vasoconstriction appears to play a key role in chronic PHT in our two neonatal rat models. Inhibitors of ROCK have potential as a testable therapy in neonates with PHT that is refractory to NO.

scholarly journals Microparticulate/nanoparticulate powders of a novel Nrf2 activator and an aerosol performance enhancer for pulmonary delivery targeting the lung Nrf2/Keap-1 pathway

2016 ◽  
Vol 1 (1) ◽  
pp. 48-65 ◽  
Author(s):  
Priya Muralidharan ◽  
Don Hayes ◽  
Stephen M. Black ◽  
Heidi M. Mansour
Keyword(s):  
Solid State ◽  
Engineering Design ◽  
Physicochemical Characterization ◽  
Pulmonary Delivery ◽  
Dry Powder Inhalers ◽  
Particle Engineering ◽  
Aerosol Performance ◽  
Nrf2 Activator ◽  
Respirable Particle

Solid-state respirable particle engineering design, physicochemical characterization, & in vitro aerosolization of advanced microparticulate/nanoparticulate dry powder inhalers targeting the lung Nrf2/Keap-1 pathway.

scholarly journals Rescue treatment with a Rho-kinase inhibitor normalizes right ventricular function and reverses remodeling in juvenile rats with chronic pulmonary hypertension

2010 ◽  
Vol 299 (6) ◽  
pp. H1854-H1864 ◽  
Author(s):  
Emily Z. Xu ◽  
Crystal Kantores ◽  
Julijana Ivanovska ◽  
Doreen Engelberts ◽  
Brian P. Kavanagh ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Smooth Muscle ◽  
Right Ventricular ◽  
Systolic Function ◽  
Rho Kinase ◽  
Rock Inhibitor ◽  
Rescue Treatment ◽  
Right Ventricular Systolic Function ◽  
Chronic Pulmonary Hypertension ◽  
Pulmonary Arterial

Chronic pulmonary hypertension in infancy and childhood is characterized by a fixed and progressive increase in pulmonary arterial pressure and resistance, pulmonary arterial remodeling, and right ventricular hypertrophy and systolic dysfunction. These abnormalities are replicated in neonatal rats chronically exposed to hypoxia from birth in which increased activity of Rho-kinase (ROCK) is critical to injury, as evidenced by preventive effects of ROCK inhibitors. Our objective in the present study was to examine the reversing effects of a late or rescue approach to treatment with a ROCK inhibitor on the pulmonary and cardiac manifestations of established chronic hypoxic pulmonary hypertension. Rat pups were exposed to air or hypoxia (13% O2) from postnatal day 1 and were treated with Y-27632 (15 mg/kg) or saline vehicle by twice daily subcutaneous injection commencing on day 14, for up to 7 days. Treatment with Y-27632 significantly attenuated right ventricular hypertrophy, reversed arterial wall remodeling, and completely normalized right ventricular systolic function in hypoxia-exposed animals. Reversal of arterial wall remodeling was accompanied by increased apoptosis and attenuated content of endothelin (ET)-1 and ETA receptors. Treatment of primary cultured juvenile rat pulmonary artery smooth muscle cells with Y-27632 attenuated serum-stimulated ROCK activity and proliferation and increased apoptosis. Smooth muscle apoptosis was also induced by short interfering RNA-mediated knockdown of ROCK-II, but not of ROCK-I. We conclude that sustained rescue treatment with a ROCK inhibitor reversed both the hemodynamic and structural abnormalities of chronic hypoxic pulmonary hypertension in juvenile rats and normalized right ventricular systolic function. Attenuated expression and activity of ET-1 and its A-type receptor on pulmonary arterial smooth muscle was a likely contributor to the stimulatory effects of ROCK inhibition on apoptosis. In addition, our data suggest that ROCK-II may be dominant in enhancing survival of pulmonary arterial smooth muscle.

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