Effect Of Carbonic Anhydrase Inhibition Research Articles

Novel Drug Screening Assay for Acanthamoeba castellanii and the Anti-Amoebic Effect of Carbonic Anhydrase Inhibitors.

Acanthamoeba castellanii is an amoeba that inhabits soil and water in every part of the world. Acanthamoeba infection of the eye causes keratitis and can lead to a loss of vision. Current treatment options are only moderately effective, have multiple harmful side effects, and are tedious. In our study, we developed a novel drug screening method to define the inhibitory properties of potential new drugs against A. castellanii in vitro. We found that the clinically used carbonic anhydrase inhibitors, acetazolamide, ethoxzolamide, and dorzolamide, have promising antiamoebic properties.

Amyloid Beta Oligomers Activate Death Receptors and Mitochondria-Mediated Apoptotic Pathways in Cerebral Vascular Smooth Muscle Cells; Protective Effects of Carbonic Anhydrase Inhibitors.

Amyloid beta (Aβ) deposition within the brain vasculature is an early hallmark of Alzheimer's disease (AD), which triggers loss of brain vascular smooth muscle cells (BVSMCs) in cerebral arteries, via poorly understood mechanisms, altering cerebral blood flow, brain waste clearance, and promoting cognitive impairment. We have previously shown that, in brain endothelial cells (ECs), vasculotropic Aβ species induce apoptosis through death receptors (DRs) DR4 and DR5 and mitochondria-mediated mechanisms, while FDA-approved carbonic anhydrase inhibitors (CAIs) prevent mitochondria-mediated EC apoptosis in vitro and in vivo. In this study, we analyzed Aβ-induced extrinsic and intrinsic (DR- and mitochondria-mediated) apoptotic pathways in BVSMC, aiming to unveil new therapeutic targets to prevent BVSMC stress and death. We show that both apoptotic pathways are activated in BVSMCs by oligomeric Aβ42 and Aβ40-Q22 (AβQ22) and mitochondrial respiration is severely impaired. Importantly, the CAIs methazolamide (MTZ) and acetazolamide (ATZ) prevent the pro-apoptotic effects in BVSMCs, while reducing caspase 3 activation and Aβ deposition in the arterial walls of TgSwDI animals, a murine model of cerebral amyloid angiopathy (CAA). This study reveals new molecular targets and a promising therapeutic strategy against BVSMC dysfunction in AD, CAA, and ARIA (amyloid-related imaging abnormalities) complications of recently FDA-approved anti-Aβ antibodies.

Prospects for the application of inhibitors of carbonic anhydrase isoforms IX and XII in oncology

Human carbonic anhydrase isoforms IX and XII play a key role in maintaining acid-base balance in solid tumors, creating a favorable microenvironment for the growth, invasion and metastasis of tumor cells. In the last few years, a number of scientific groups have published results that inhibition of isoforms IX and XII significantly increases the effectiveness of classical chemotherapy, makes it possible to suppress the resistance of tumor cells to chemotherapy and increase their sensitivity to the used drugs (including reducing the dose of cytostatics). In the review, we analyzed the scientific literature on the role of carbonic anhydrase isoforms IX and XII in carcinogenesis and on the combined effect of carbonic anhydrase inhibitors with antitumor drugs.

Membrane Permeability Is Required for the Vasodilatory Effect of Carbonic Anhydrase Inhibitors in Porcine Retinal Arteries.

It has been demonstrated previously that a variety of carbonic anhydrase inhibitors (CAIs) can induce vasodilation in pre-contracted retinal arteriolar segments although with different efficacy and potency. Since the CAIs tested so far are able to permeate cell membranes and inhibit both intracellular and extracellular isoforms of the enzyme, it is not clear whether extra- or intracellular isoforms or mechanisms are mediating their vasodilatory effects. By means of small wire myography, we have tested the effects of four new CAIs on wall tension in pre-contracted retinal arteriolar segments that demonstrably do not enter cell membranes but have high affinity to both cytosolic and membrane-bound isoforms of CA. At concentrations between 10-6 M to 10-3 M, none of the four membrane impermeant CAIs had any significant effect on arteriolar wall tension, while the membrane permeant CAI benzolamide (10-3 M) fully dilated all arteriolar segments tested. This suggests that CAI act as vasodilators through cellular mechanisms located in the cytoplasm of vascular cells.

Neuroprotective Effects of Carbonic Anhydrase Inhibition and Cyclic Adenosine Monophosphate Activation in Mouse Model of Transient Global Cerebral Ischemia and Reperfusion.

Cerebral ischemia is the primary basis of stroke, both sharing common pathogenic origins leading to irreversible brain damage if blood supply is not restored promptly. Existing evidence indicates that carbonic anhydrase (CA) inhibitors (CAIs) may impart therapeutic benefits against ischemia-reperfusion (I/R) pathology via the adenylyl cyclase-cyclic adenosine monophosphate (cAMP) pathway. We hypothesize that CAI and cAMP activation may enhance the therapeutic outcome against I/R conditions. In this investigation, the potential of dichlorphenamide (CAI) and the role of cAMP against ischemia-reperfusion injury were evaluated using a transient global cerebral I/R (tGCI/R) model. Swiss albino mice were subjected to bilateral common carotid artery occlusion (BCCAo) for 20min and reperfusion (R) or sham surgery on day 1. Dichlorphenamide (DCPA, 20mg/kg) and/or forskolin (cAMP agonist, 3mg/kg) was administered intraperitoneally (i.p.) after BCCAo/R for 14days. Results showed that tGCI/R impaired neurocognitive functions and lowered brain levels of cAMP and protein kinase A (PKA) that were ameliorated by DCPA and/or forskolin (FSK). DCPA and/or FSK attenuated tGCI/R-induced brain edema, blood-brain barrier dysfunction, oxidative-nitrosative stress, pro-inflammatory cytokines, acetylcholinesterase activity, cell death, and neurotransmitter imbalance (e.g., glutamate, γ-aminobutyric acid). The study showed that DCPA improved neurological and biochemical parameters against tGCI/R injury via cAMP-PKA-mediated activation of protective mechanisms. However, DCPA and FSK in combination showed much enhanced therapeutic outcomes against tGCI/R. Therefore, CA and cAMP present novel targets that may retard the progress of a transient ischemic attack to a full-blown stroke.

The gram-negative bacterium Escherichia coli as a model for testing the effect of carbonic anhydrase inhibition on bacterial growth

Carbonic anhydrases, catalysing the reversible CO2 hydration reaction, contribute in all living organisms to the maintenance of stable metabolic functions depending on intracellular concentrations of carbon dioxide, bicarbonate, and protons. Recent studies have examined how CAs affect bacterial lifecycle, considering these enzymes druggable targets due to interference with their activities by using inhibitors or activators. Here, we propose Escherichia coli cells as a model for testing the effect of acetazolamide (AZA), a potent CA inhibitor, on bacterial survival by evaluating E. coli growth through its glucose consumption. AZA, at concentrations higher than 31.2 µg/mL, was able to impair E. coli growth and glucose uptake. AZA is a good inhibitor of the two recombinant E. coli CAs, the β-CA CynT2, and the γ-CA EcoCAγ, with KIs of 227 and 248 nM, respectively. This study provides a proof-of-concept, low-cost method for identifying effective CA inhibitors capable of impairing bacterial metabolism.

Anticonvulsant Effects of Carbonic Anhydrase Inhibitors: The Enigmatic Link Between Carbonic Anhydrases and Electrical Activity of the Brain.

Acetazolamide (ACZ), a sulfonamide carbonic anhydrase (CA) inhibitor, was first introduced into medical use as a diuretic in the1950s. Shortly after its introduction, its antiglaucoma and anticonvulsant properties came to light. Subsequently, studies of ACZ have explored a plethora of neurophysiological functions of CAs in the CNS. In addition, topiramate (TPM) and zonisamide (ZNS), which were developed as antiepileptic drugs (AEDs) in the1990s, were found to have the ability to inhibit CAs. How CA inhibition prevents seizures is elusive. CA expression and activity are extensively detected in neurons, the choroid plexus, oligodendrocytes and astrocytes. TPM and ZNS appear to produce multimodal actions in the CNS as well as CA inhibition unlike ACZ. Nonetheless, CA inhibitors share some common denominators. They do not only affect the fine equilibrium among CO2, H+ and HCO3- in the extraneuronal and intraneuronal milieu, but also modulate the activity of ligand gated ion channels at the neuronal level such as GABA-A signaling through inhibiting CA-replenished HCO3- efflux. In addition, there are studies reporting their ability to alter Ca2+ kinetics through modulation of ligand gated Ca2+ channels, voltage gated Ca2+ channels (VGCC) or Ca2+-induced Ca2+ release channels (CICRC). The present study will review the involvement of CAs in the formation of epileptogenesis, and likely mechanisms by which CA inhibitors suppress the electrical activity of the brain. The common properties of CA inhibitors provide some clues for a possible link among metabolism, CAs, Ca2+ and GABA signaling.

The effects of cardiac drugs on human erythrocyte carbonic anhydrase I and II isozymes

Cardiovascular diseases are the leading cause of mortality worldwide. In recent years, the relationship between carbonic anhydrase inhibitors and atherosclerosis has attracted attention. In this study, we aimed to determine the in vitro effects of 35 frequently used cardiac drugs on human carbonic anhydrase I (hCA I) and II (hCA II). The inhibitory effects of the drugs on hCA I and hCA II were determined with both the hydratase and esterase methods. The most potent inhibitors observed were propafenone (hCA I: 2.8 µM and hCA II: 3.02 µM) and captopril (hCA I: 1.58 µM and hCA II: 6.25 µM). Isosorbide mononitrate, propranolol, furosemide, and atorvastatin were also potent inhibitors. The inhibitor constant, K i, value from the Lineweaver–Burk plot for propafenone was 2.38 µM for hCA I and 2.97 µM for hCA II. The tested cardiac drugs showed potent in vitro inhibition of the hCA I and II isozymes. Especially, in patients with atherosclerotic heart disease, these drugs may be preferred primarily due to the beneficial effects of carbonic anhydrase inhibition on atherosclerosis.

Carbonic Anhydrase Inhibition Ameliorates Inflammation and Experimental Pulmonary Hypertension.

Inflammation and vascular smooth muscle cell (VSMC) phenotypic switching are causally linked to pulmonary arterial hypertension (PAH) pathogenesis. Carbonic anhydrase inhibition induces mild metabolic acidosis and exerts protective effects in hypoxic pulmonary hypertension. Carbonic anhydrases and metabolic acidosis are further known to modulate immune cell activation. To evaluate if carbonic anhydrase inhibition modulates macrophage activation, inflammation, and VSMC phenotypic switching in severe experimental pulmonary hypertension, pulmonary hypertension was assessed in Sugen 5416/hypoxia (SU/Hx) rats after treatment with acetazolamide or ammonium chloride (NH4Cl). We evaluated pulmonary and systemic inflammation and characterized the effect of carbonic anhydrase inhibition and metabolic acidosis in alveolar macrophages and bone marrow-derived macrophages (BMDMs). We further evaluated the treatment effects on VSMC phenotypic switching in pulmonary arteries and pulmonary artery smooth muscle cells (PASMCs) and corroborated some of our findings in lungs and pulmonary arteries of patients with PAH. Both patients with idiopathic PAH and SU/Hx rats had increased expression of lung inflammatory markers and signs of PASMC dedifferentiation in pulmonary arteries. Acetazolamide and NH4Cl ameliorated SU/Hx-induced pulmonary hypertension and blunted pulmonary and systemic inflammation. Expression of carbonic anhydrase isoform 2 was increased in alveolar macrophages from SU/Hx animals, classically (M1) and alternatively (M2) activated BMDMs, and lungs of patients with PAH. Carbonic anhydrase inhibition and acidosis had distinct effects on M1 and M2 markers in BMDMs. Inflammatory cytokines drove PASMC dedifferentiation, and this was inhibited by acetazolamide and acidosis. The protective antiinflammatory effect of acetazolamide in pulmonary hypertension is mediated by a dual mechanism of macrophage carbonic anhydrase inhibition and systemic metabolic acidosis.

Effects of carbonic anhydrase inhibition on biomass and primary production of estuarine benthic microalgal communities

Recent studies have focused on carbon concentrating mechanisms and the associated enzyme, carbonic anhydrase, to better understand the efficiency of CO2 uptake rates and carbon fixation in photoautotrophs. Some benthic microalgae (BMA) may be limited by inorganic carbon availability because high photosynthetic rates withdraw a large amount of CO2 and HCO3– in the top layer of sediment. Investigating the mechanisms that affect carbon acquisition are necessary if we are to fully understand the functioning and structuring processes of these systems. From this, we can better predict the potential impacts of increasing atmospheric CO2 concentrations on BMA communities. The purpose of this research was to examine a carbon concentrating mechanism used by BMA through their responses to induced carbon limitation. This approach was conducted through the removal of carbonic anhydrase (CA) activity using an inhibitor, ethoxyzolamide. Microcosm experiments were performed on intertidal muddy sediments from North Inlet Estuary, SC. Exposure to ethoxyzolamide resulted in a reduction of gross primary productivity (GPP) without a reduction in total BMA biomass. Furthermore, removed CA activity caused BMA cumulative GPP maxima to shift upward toward the surface in the sediment column. Active CA was necessary to maintain high GPP rates in these communities and allowed motile BMA to use a wider portion of the sediment column. Available HCO3– at lower depths could still be dehydrated into CO2 by microalgae with CA. Changes in global atmospheric CO2 concentrations leading to higher CO2 availability at the atmosphere-sediment interface may alter the structure and function of these BMA systems, and the vertical distribution of GPP. These consequences may have important implications for the biogeochemical cycling occurring in estuaries.

Correlation of carbonic anhydrase and Rubisco in the growth and photosynthesis in the diatom Phaeodactylum tricornutum

Carbonic anhydrase (CA) and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) are critical enzymes involved in the CO2-concentrating mechanism (CCM) and carbon fixation in phytoplankton. These enzymes play key roles in photosynthetic carbon acquisition and fixation in diatoms. However, the potential synergistic interactions of the said enzymes remain little understood. In the present study, effects of CA inhibitors on the growth, photosynthetic rate, and Rubisco activities in the diatom Phaeodactylum tricornutum were investigated. Our results showed that the growth and photosynthetic O2 evolution rate of P. tricornutum were significantly inhibited by the intracellular CA (iCA) inhibitor ethoxzolamide (EZ), but not by extracellular CA (eCA) inhibitor acetazolamide (AZ) in the long term. This suggests that the eCA was not necessarily essential for growth and photosynthesis, but rather it was the iCA activity that was more crucial in P. tricornutum. We also found that there is a significant positive correlation between Rubisco and CA activities, suggesting that the two carbon acquisition and fixation components of photosynthesis may have undergone co-evolution to optimize fundamental trade-offs. These results further indicate that the roles of eCA and iCA may also differ among diatom species or strains, and that their functions are tightly correlated with other carbon fixation components.

Acetazolamide Reduces Blood Pressure and Sleep-Disordered Breathing in Patients With Hypertension and Obstructive Sleep Apnea: A Randomized Controlled Trial.

The carbonic anhydrase inhibitor acetazolamide (AZT) modulates blood pressure at high altitude and reduces sleep-disordered breathing in patients with obstructive sleep apnea (OSA). We aimed to investigate the treatment effect of AZT and in combination with continuous positive airway pressure (CPAP) on blood pressure in patients with hypertension and OSA. In a prospective, randomized, three-way crossover study, 13 male patients with hypertension and moderate to severe OSA (age 64 ± 7 years, body mass index 29 ± 4 kg/m2, and mean apnea-hypopnea index 37 ± 23 events/h) received AZT, CPAP, or AZT plus CPAP for 2-week periods. Antihypertensive medication was washed out. Office and 24-hour blood pressure, arterial stiffness, polygraphic sleep study data, and blood chemistry were compared. AZT alone and AZT plus CPAP, but not CPAP alone, reduced office mean arterial pressure compared to baseline (-7 [95% CI -11 to -4], -7 [95% CI -11 to -4] and -1 [95% CI -5 to 4] mmHg, respectively; repeated- measures analysis of variance (RM-ANOVA; P = .015). Aortic systolic pressure and augmentation index, assessed by radial artery oscillatory tonometry, were unaffected by CPAP but decreased after AZT and AZT plus CPAP (RM-ANOVA P = .030 and .031, respectively). The apnea-hypopnea index was significantly reduced in all three treatment arms, most prominently by AZT plus CPAP (RM-ANOVA P = .003). The reduction of venous bicarbonate concentration following AZT was correlated with the change of apnea-hypopnea index (r = 0.66, P = .013). AZT reduced blood pressure, vascular stiffness, and sleep-disordered breathing in patients with OSA and comorbid hypertension. Carbonic anhydrase inhibition may constitute a potential target for drug therapy in patients with sleep apnea and comorbid hypertension. Registry: ClinicalTrials.gov; Identifier: NCT02220803; Title: A Short Term Open, Randomized Cross-over Trial Exploring the Effect of Carbonic Anhydrase Inhibition by Acetazolamide on Sleep Apnea Associated Hypertension and Vascular Dysfunction; URL: https://clinicaltrials.gov/ct2/show/NCT02220803 and Registry: EU Clinical Trials Register; EudraCT Number: 2013-004866-33; Title: A short term open, randomized cross over trial exploring the effect of carbonic anhydrase inhibition by acetazolamide on sleep apnea associated hypertension; URL: https://www.clinicaltrialsregister.eu/ctr-search/search?query=2013-004866-33.

Carbonic Anhydrase Inhibitors for the Treatment of Cystic Macular Lesions in Children With X-Linked Juvenile Retinoschisis.

Little is known regarding the therapeutic effect of carbonic anhydrase inhibitors (CAIs) in the management of cystic macular lesions in children with X-linked juvenile retinoschisis (XLRS) despite the fact that this disease often manifests during childhood. Therefore, our goal was to determine the efficacy of CAIs in the treatment of cystic macular lesions in children with XLRS. We used CAIs to treat cystic macular lesions in 18 eyes of nine children with XLRS. We evaluated the therapeutic effect of CAI treatment with the best-corrected visual acuity and foveal zone thickness (FZT) with spectral-domain optical coherence tomography. A reduction of at least 22.4% in FZT was defined as objective evidence of response. Five of nine (55.6%) XLRS patients showed a significant reduction of FZT in both eyes over a median treatment interval of 6.8 months (range, 1-23). In four of five (80.0%) patients, this reduction was already apparent after 1 month of treatment. An improvement of visual acuity was observed in five eyes (27.8%) of three patients (33.3%). Six patients (66.6%) reported minor side effects. Treatment with CAIs decreased FZT in more than half of the children with XLRS. This effect was observed within 1 month in the majority of patients. Carbonic anhydrase inhibitor treatment restores retinal anatomy and may contribute to creating optimal circumstances for gene therapy.

Effect of carbonic anhydrase inhibitors on Anopheles albimanus larvae (Diptera: Culicidae)

Carbonic anhydrase (CA) catalyzes the reversible conversion of CO2 to bicarbonate and participates in mechanisms of alkalinization in the intestine of mosquitoes. The toxicity of four CA inhibitors (CAI ): acetazolamide (AZ M), methazolamide (MZM), ethoxolamide (ETX) and dorzolamide (DZA ) were evaluated in larvae of Anopheles albimanus by monitoring mortality 24, 48 and 72 hours post application, at a concentration of 50 ug/ml diluted in dimethyl sulfoxide previously. All IA C reduced the population of larvae in variable proportions. ETX showed the highest toxicity, achieving more than 80% mortality after 24 hours and 98% after 72 hours of application. The CAI , AZ M, MZM and DZA showed less toxicity (<50% mortality). Our results indicate that the CAI , including ETX in particular, is a worthy candidate as an alternative for the control of An. albimanus, which is considered a primary vector of malaria in Colombia.

In vivo imaging reveals novel aspects of retinal disease progression in Rs1h−/Y mice but no therapeutic effect of carbonic anhydrase inhibition

X-linked juvenile retinoschisis (XLRS) is the most common juvenile maculopathy in men and is caused by mutations in the gene encoding retinoschisin (RS1). Evidence in the literature on the therapeutic effect of carboanhydrase inhibitors (CAIs) to treat schisis formation in the retina has remained equivocal. Here, we evaluate the effect of the CAI dorzolamide on the structural and functional disease progression in the mouse model for XLRS (Rs1h(-/y)). Rs1h (-/y) mice were treated unilaterally with dorzolamide eye drops (Trusopt(®) 20 mg/mL) every 12 h for 2 weeks starting on postnatal day 14 (n = 27). Changes of retinal structure were monitored by confocal scanning laser ophthalmoscopy and spectral domain optical coherence tomography 12 h, 14 days, 4 weeks, 2 months, and 6 months after completion of the treatment. Schisis formation (peak at 3 months) preceded photoreceptor degeneration and hyper-fluorescence (peak at 7 months). Structural pathology was most severe in the superior hemi-retina with previously unreported hyper-fluorescent lesions. Quantitative analysis showed no significant differences regarding the inner or outer retinal thickness of the treated vs. untreated eyes 12 h after the completion of treatment (IRT(12 h) = -1.29 ± 1.89 μm; ORT(12 h) = 0.61 ± 2.08 μm; mean ± 95%CI) or at any later time point. Time line analysis after short-term treatment with CAI failed to show short-, intermediate-, or long-term evidence of structural improvement in Rs1h(-/y) mice. Schisis formation in the inner retina peaked at the age of 3 months and was followed by photoreceptor degeneration predominantly in the superior hemi-retina. Previously unreported hyper-fluorescent lesions co-register with structural retinal pathologies.

Effects of carbonic anhydrase inhibitors on pseudophakic corneal edema

Acetazolamide as inhibitor of carbonic anhydrase, in glaucoma therapy, has an effect on lowering intraocular pressure. In addition to this primary effects, we attempted to determine the effect on the cornea after phacoemulsification. In our study, we determined the effects of systemically applied dose of 500 mg of acetazolamide on the edema, thickness, and corneal morphology after phacoemulsification ,with the Anterior Segment Optical Coherence Tomography (AS OCT). The study included 53 patients who were divided into two groups. Groups were stratified by type of cataract (nuclear), age (62+/-1.5), preoperative visual acuity (5/60) Snellen table,and preoperative findings on AS-OCT. Phacoemulsification has done the same surgeon, with the same ultrasound probe length using ultrasonic force of the average value of 14 for a period of 72+/-2.5 seconds. Group of 33 patients were administered systemic acetazolamide at a dose of 250 mg per scheme 01 hours +24 hours after phacoemulsification (group I). Another group of 20 patients did not receive acetazolamide(group II). AS OCT recordings were performed before surgery, 6 hours and 24 hours after surgery. The mean value in both groups was 549+/-9 microm before surgery. Group I had average value of 648+/-6 microm after 6h, and the mean value was 612+/-4 microm after 24h. The group II,had a mean thickness of the cornea after 6 hours of 720+/-5 microm, and after 24 hours 708+/-4 microm. Morphological changes in the tomograms of the group I showed minimal creases Descemet's membrane. Postoperative visual acuity was 0,6,24h after the surgery in the I group of patients,and 0,3 in the II group. In our study the patients who administered systemic acetazolamide had significant reduction of central corneal thickness. The folds of Descemet's membrane and endothelial dysfunction in AS OCT tomograms showed less structural changes in group I. Significant better postoperative visual acuity in these patients is probably because of the smaller corneal edema after 24 h, which improves patients' comfort.

Influence of topical carbonic anhydrase inhibitor on the expression of aquaporin-1 in rat cornea with neovascularization

Background Researches showed that aquaporin-1 (AQP1) is closely associated with corneal neovescularization(CNV).Carbonic anhydrase inhibitor has the inhibitory effect on the AQP1 and further suppresses the CNV.However,the systemic adverse effect of Carbonic anhydrase inhibitor limit its clinical application.Therefore,the influence of topical carbonic anhydrase inhibitor on CNV is concerned.Objective Present study was to investigate the effects of topical carbonic anhydrase inhibitors on the expression of AQP1 in rat cornea after alkali burn and explore its role in corneal neovascularization (CNV).Methods The alkali-burn animal models were established in 60 eyes of 30 clean Sprague Dawley rats by putting the filter paper soaked 1 mol/L NaOH solution at the central cornea for 40 seconds.1％ Brinzolamide was topically administered in the 30 eyes of 15 models (Brinzolamide group),and the normal saline solution was used at the same way in other 30 eyes of 15 rats (model group).The 10 eyes of 5 normal Sprague Dawley received the eye drops of normal saline solution as the normal control group.The corneal burning degree was graded on the Mahoney ' s criteria in the third day,and Ee ' s method was used to score the opacification of cornea and the CNV area was analyzed in 3,5,7,10 days under the slit lamp microscope.The cornea tissue was obtained in the tenth day after burning for the observation of the pathology under the light microscope and the ultrastructure under the transmission electron microscope.The expressions of AQP1 and vascular endothelial growth factor(VEGF) in cornea tissue were detected using immunohistochemistry.The use of animals complied with the Statement of ARVO.Results No significant difference was seen in the scores of rat corneal alkali burn between the model group and brinzolamide group( t=0.97,P＞0.05 ).The scores of corneal edema and opacification and neovascular area were lower in brinzolamide group compared with model group ( t =2.18,P＜0.05 ;t =6.58,P＜0.01 ).The pathological and ultrastructural examinations showed less CNV and inflammatory cells in rat corneal tissue of the brinzolamide group in comparison with model group.The grey values of VEGF were 84.92±9.49 and 78.18± 11.41,and those of AQP1 were 88.01 ± 11.03 and 58.10 ± 12.14 in the model group and brinzolamidegroup respectively,showing statistically significant differences ( VEGF:t =2.48,P =0.02 ; AQPI:t =9.99,P =0.00 ).Conclusions 1％ Brinzolamide suppresses alkali burn-induced CNV by downregulating the expressions of AQP1 and VEGF in cornea in rat. Key words: Topical carbonic anhydrase inhibitor; Brinzolamide ; Corneal neovascularization; Aquaporin-1 ; Vascular endothelial growth factor

Extremely low-frequency electromagnetic fields affect lipid-linked Carbonic anhydrase

In the last years, the effect of extremely low-frequency electromagnetic fields (ELF-EMF) on the activity of different enzymes were investigated. Only the membrane-anchored enzymes did decrease their activity, up to 50%. In this work, the effect of ELF-EMF on bovine lung membrane carbonic anhydrase (CA) were studied. Carbonic anhydrases are a family of 14 zinc-containing isozymes catalyzing the reversible reaction: CO2+H2O = HCO3− +H+. CA differ in catalytic activity and subcellular localization. CA IV, IX, XII, XIV, and XV are membrane bound. In particular, CA IV, which is expressed in the lung, is glycosyl phosphatidyl inositol-linked to the membrane, therefore it was a candidate to inhibition by ELF-EMF. Exposure to the membranes to a field of 75 Hz frequency and different amplitudes caused CA activity to a reproducible decrease in enzymatic activity by 17% with a threshold of about 0.74 mT. The decrease in enzymatic activity was independent of the time of permanence in the field and was completely reversible. When the source of enzyme was solubilized with Triton, the field lost its effect on CA enzymatic activity, suggesting a crucial role of the membrane, as well as of the particular linkage of the enzyme to it, in determining the conditions for CA inactivation. Results are discussed in terms of the possible physiologic effects of CA inhibition in target organs.

Effects of chronic acetazolamide administration on gas exchange and acid‐base control in pulmonary circulation in exercising horses

Carbonic anhydrase (CA) catalyses the hydration/dehydration reaction of CO(2) and increases the rate of Cl(-) and HCO(3)(-) exchange between the erythrocytes and plasma. Therefore, chronic inhibition of CA has a potential to attenuate CO(2) output and induce greater metabolic and respiratory acidosis in exercising horses. To determine the effects of Carbonic anhydrase inhibition on CO(2) output and ionic exchange between erythrocytes and plasma and their influence on acid-base balance in the pulmonary circulation (across the lung) in exercising horses with and without CA inhibition. Six horses were exercised to exhaustion on a treadmill without (Con) and with CA inhibition (AczTr). CA inhibition was achieved with administration of acetazolamide (10 mg/kg bwt t.i.d. for 3 days and 30 mg/kg bwt before exercise). Arterial, mixed venous blood and CO(2) output were sampled at rest and during exercise. An integrated physicochemical systems approach was used to describe acid base changes. AczTr decreased the duration of exercise by 45% (P < 0.0001). During the transition from rest to exercise CO(2) output was lower in AczTr (P < 0.0001). Arterial PCO(2) (P < 0.0001; mean ± s.e. 71 ± 2 mmHg AczTr, 46 ± 2 mmHg Con) was higher, whereas hydrogen ion (P = 0.01; 12.8 ± 0.6 nEq/l AczTr, 15.5 ± 0.6 nEq/l Con) and bicarbonate (P = 0.007; 5.5 ± 0.7 mEq/l AczTr, 10.1 ± 1.3 mEq/l Con) differences across the lung were lower in AczTr compared to Con. No difference was observed in weak electrolytes across the lung. Strong ion difference across the lung was lower in AczTr (P = 0.0003; 4.9 ± 0.8 mEq AczTr, 7.5 ± 1.2 mEq Con), which was affected by strong ion changes across the lung with exception of lactate. CO(2) and chloride changes in erythrocytes across the lung seem to be the major contributors to acid-base and ions balance in pulmonary circulation in exercising horses.

Chlorthalidone: Don’t Call It “Thiazide-Like” Anymore

Recently, there has been growing interest in the idea that chlorthalidone should be preferred over thiazides when it comes to selecting a nonloop diuretic for treatment of hypertension.1–4 Depending on how these drugs are dosed, it is believed that the nonthiazide diuretic chlorthalidone might provide greater cardiovascular protection than thiazide diuretics because of its more potent and prolonged effects on blood pressure. However, given that the chemical structure of chlorthalidone is clearly distinct from the basic chemical structure of the thiazides (Figure), it is possible that the nonthiazide chlorthalidone might also differ from thiazides with respect to effects on determinants of cardiovascular disease other than just blood pressure. Yet, few if any studies have ever directly compared the effects of the nonthiazide chlorthalidone versus thiazides on potential cellular or molecular mechanisms of cardiovascular morbidity and mortality independent of blood pressure. Figure. The nonthiazide diuretic chlorthalidone and the thiazides, including hydrochlorothiazide, have sulfonamide groups (denoted within the unshaded ovals) that interact with the Zn(II) ion in the active site cleft of CA and with amino acid residues in the enzyme as well. A sulfonamide moiety is also present in the nonthiazide diuretics metolazone and indapamide (not shown), and these agents are also known to affect CA activity. However, the nonthiazide diuretics chlorthalidone, metolazone, and indapamide do not have the benzothiadiazine dioxide scaffold that is characteristic of the thiazides (denoted within the shaded oval). The structures of the nonthiazide diuretics beyond the sulfonamide group are clearly different from those of the thiazides and contribute to different effects of the various diuretics on CA isozyme activity profiles and perhaps to different effects on cardiovascular outcomes as well. In this issue of Hypertension , Woodman et al5 report novel in vitro experiments in which chlorthalidone was found to reduce epinephrine-induced platelet aggregation …