Cryo-EM reveals mechanisms of angiotensin I-converting enzyme allostery and dimerization.

Hypertension (high blood pressure) is a major risk factor for cardiovascular disease, which is the leading cause of death worldwide. The somatic isoform of angiotensin I-converting enzyme (sACE) plays a critical role in blood pressure regulation, and ACE inhibitors are thus widely used to treat hypertension and cardiovascular disease. Our current understanding of sACE structure, dynamics, function, and inhibition has been limited because truncated, minimally glycosylated forms of sACE are typically used for X-ray crystallography and molecular dynamics simulations. Here, we report the first cryo-EM structures of full-length, glycosylated, soluble sACE (sACES1211 ). Both monomeric and dimeric forms of the highly flexible apo enzyme were reconstructed from a single dataset. The N- and C-terminal domains of monomeric sACES1211 were resolved at 3.7 and 4.1 Å, respectively, while the interacting N-terminal domains responsible for dimer formation were resolved at 3.8 Å. Mechanisms are proposed for intradomain hinging, cooperativity, and homodimerization. Furthermore, the observation that both domains were in the open conformation has implications for the design of sACE modulators.

Renal angiotensin I-converting enzyme-deficient mice are protected against aristolochic acid nephropathy.

The renal renin-angiotensin system (RAS) is involved in the development of chronic kidney disease. Here, we investigated whether mice with reduced renal angiotensin I-converting enzyme (ACE-/-) are protected against aristolochic acid nephropathy (AAN). To further elucidate potential molecular mechanisms, we assessed the renal abundances of several major RAS components. AAN was induced using aristolochic acid I (AAI). Glomerular filtration rate (GFR) was determined using inulin clearance and renal protein abundances of renin, angiotensinogen, angiotensin I-converting enzyme (ACE) 2, and Mas receptor (Mas) were determined in ACE-/- and C57BL/6J control mice by Western blot analyses. Renal ACE activity was determined using a colorimetric assay and renal angiotensin (Ang) (1-7) concentration was determined by ELISA. GFR was similar in vehicle-treated mice of both strains. AAI decreased GFR in controls but not in ACE-/- mice. Furthermore, AAI decreased renal ACE activity in controls but not in ACE-/- mice. Vehicle-treated ACE-/- mice had significantly higher renal ACE2 and Mas protein abundances than controls. AAI decreased renal ACE2 protein abundance in both strains. Furthermore, AAI increased renal Mas protein abundance, although the latter effect did not reach statistical significance in the ACE-/- mice. Renal Ang(1-7) concentration was similar in vehicle-treated mice of both strains. AAI increased renal Ang(1-7) concentration in the ACE-/- mice but not in the controls. Mice with reduced renal ACE are protected against AAN. Our data suggest that in the face of renal ACE deficiency, AAI may activate the ACE2/Ang(1-7)/Mas axis, which in turn may deploy its reno-protective effects.

Angiotensin-converting enzymes as druggable features of psychiatric and neurodegenerative disorders.

The renin-angiotensin system (RAS) plays essential roles in maintaining peripheral cardiovascular homeostasis, with its potential roles in the brain only being recognized more recently. Angiotensin-I-converting enzyme (ACE) is the main component of the RAS, and it has been implicated in various disorders of the brain. ACE and other RAS components, including the related enzyme ACE2, angiotensin peptides and their respective receptors, can participate in the pathological state, as well as with potential to contribute to neuroprotection and/or to complement existing treatments for psychiatric illness. In this narrative review, we aimed to identify the main studies describing the functions of the RAS and ACEs in the brain and their association with brain disorders. These include neurodegenerative disorders such as Parkinson's and Alzheimer's diseases, psychiatric illnesses such as schizophrenia, bipolar disorder, and depression. We also discuss the possible association of a functional polymorphism of the ACE gene with these brain diseases and the relevance of the neuroprotective and anti-inflammatory properties of ACE inhibitors (ACEis) and angiotensin receptor blockers (ARBs). Based on this, we conclude that there is significant potential value to the inclusion of ACEis and/or ARBs as a novel integrated approach for the treatment of various disorders of the brain, and particularly for psychiatric illness.

Evaluation of a biotin-based surrogate virus neutralization test for detecting postvaccination antibodies against SARS-CoV-2 variants in sera.

A severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) surrogate virus neutralization test (sVNT) was used to determine the degree of inhibition of binding between human angiotensin converting enzyme 2 (hACE2) and the receptor binding domain (RBD) of spike protein by neutralizing antibodies in a biosafety level 2 facility. Here, to improve the sensitivity and specificity of the commercial sVNT, we developed a new biotin based sVNT using biotinylated RBD and HRP conjugated streptavidin instead of HRP conjugated RBD for direct detection in an ELISA assay that strongly correlated to the FDA approved cPass sVNT commercial kit (R2 = 0.8521) and pseudo virus neutralization test (R2 = 0.9006) (pVNT). The biotin based sVNT was evaluated in 535 postvaccination serum samples corresponding to second and third boosts of AZD1222 and BNT162b2 vaccines of the wild type strain. We confirmed that the neutralizing antibodies against SARS-CoV-2 variants in second vaccination sera decreased after a median of 141.5 days. Furthermore, vaccination sera from BNT162b2-BNT162b2 vaccines maintained neutralizing antibodies for longer than those of AZD1222 only vaccination. In addition, both vaccines maintained high neutralizing antibodies in third vaccination sera against Omicron BA.2 after a median of 27 days, but neutralizing antibodies significantly decreased after a median of 141.5 days. Along with the cPass sVNT commercial kit, biotin based sVNTs may also be suitable for specifically detecting neutralizing antibodies against multiple SARS-CoV-2 variants; however, to initially monitor the neutralizing antibodies in vaccinated sera using high throughput screening, conventional PRNT could be replaced by sVNT to circumvent the inconvenience of a long test time.

In silico identification of novel ACE and DPP-IV inhibitory peptides derived from buffalo milk proteins and evaluation of their inhibitory mechanisms.

The capacity of buffalo milk proteins to release bioactive peptides was evaluated and novel bioactive peptides were identified. The sequential similarity between buffalo milk proteins and their cow counterparts was analysed. Buffalo milk proteins were simulated to yield theoretical peptides via in silico proteolysis. The potential of selected proteins to release specific bioactive peptides was evaluated by the A value obtained from the BIOPEP-UWM database (Minkiewicz et al. in Int J Mol Sci 20(23):5978, 2019). Buffalo milk protein is a suitable precursor to produce bioactive peptides, particularly dipeptidyl peptidase IV (DPP-IV) and angiotensin I-converting enzyme (ACE) inhibitory peptides. Two novel ACE inhibitory peptides (KPW and RGP) and four potential DPP-IV inhibitory peptides (RGP, KPW, FPK and KFTW) derived from in silico proteolysis of buffalo milk proteins were screened using different integrated bioinformatic approaches (PeptideRanker, Innovagen, peptide-cutter and molecular docking). The Lineweaver-Burk plots showed that KPW (IC50 = 136.28 ± 10.77 µM) and RGP (104.72 ± 8.37 µM) acted as a competitive inhibitor against ACE. Similarly, KFTW (IC50 = 873.92 ± 32.89 µM) was also a competitive inhibitor of DPP-IV, while KPW and FPK (82.52 ± 10.37 and 126.57 ± 8.45 µM, respectively) were mixed-type inhibitors. It should be emphasized that this study does not involve any clinical trial.

Purification and Molecular Docking Study on the Angiotensin I-Converting Enzyme (ACE)-Inhibitory Peptide Isolated from Hydrolysates of the Deep-Sea Mussel Gigantidas vrijenhoeki.

The objective of this study was to prepare an angiotensin I-converting enzyme (ACE)-inhibitory peptide from the hydrothermal vent mussel, Gigantidas vrijenhoeki. The G. vrijenhoeki protein was hydrolyzed by various hydrolytic enzymes. The peptic hydrolysate exhibited the highest ACE-inhibitory activity and was fractionated into four molecular weight ranges by ultrafiltration. The <1 kDa fraction exhibited the highest ACE inhibitory activity and was found to have 11 peptide sequences. Among the analyzed peptides, KLLWNGKM exhibited stronger ACE inhibitory activity and an IC50 value of 0.007 µM. To investigate the ACE-inhibitory activity of the analyzed peptides, a molecular docking study was performed. KLLWNGKM exhibited the highest binding energy (-1317.01 kcal/mol), which was mainly attributed to the formation of hydrogen bonds with the ACE active pockets, zinc-binding motif, and zinc ion. These results indicate that G. vrijenhoeki-derived peptides can serve as nutritional and pharmacological candidates for controlling blood pressure.

Angiotensin-I-Converting Enzyme (ACE)-Inhibitory Peptides from the Collagens of Monkfish (Lophius litulon) Swim Bladders: Isolation, Characterization, Molecular Docking Analysis and Activity Evaluation.

The objective of this study was to isolate and characterize collagen and angiotensin-I-converting enzyme (ACE)-inhibitory (ACEi) peptides from the swim bladders of monkfish (Lophius litulon). Therefore, acid-soluble collagen (ASC-M) and pepsin-soluble collagen (PSC-M) with yields of 4.27 ± 0.22% and 9.54 ± 0.51%, respectively, were extracted from monkfish swim bladders using acid and enzyme methods. The ASC-M and PSC-M contained Gly (325.2 and 314.9 residues/1000 residues, respectively) as the major amino acid, but they had low imino acid content (192.5 and 188.6 residues/1000 residues, respectively) in comparison with collagen from calf skins (CSC) (216.6 residues/1000 residues). The sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns and ultraviolet (UV) absorption spectrums of ASC-M and PSC-M illustrated that they were mainly composed of type I collagen. Subsequently, three ACEi peptides were isolated from a PSC-M hydrolysate prepared via a double-enzyme system (alcalase + neutrase) and identified as SEGPK (MHP6), FDGPY (MHP7) and SPGPW (MHP9), with molecular weights of 516.5, 597.6 and 542.6 Da, respectively. SEGPK, FDGPY and SPGPW displayed remarkable anti-ACE activity, with IC50 values of 0.63, 0.94 and 0.71 mg/mL, respectively. Additionally, a molecular docking assay demonstrated that the affinities of SEGPK, FDGPY and SPGPW with ACE were -7.3, -10.9 and -9.4 kcal/mol, respectively. The remarkable ACEi activity of SEGPK, FDGPY and SPGPW was due to their connection with the active pockets and/or sites of ACE via hydrogen bonding, hydrophobic interaction and electrostatic force. Moreover, SEGPK, FDGPY and SPGPW could protect HUVECs by controlling levels of nitric oxide (NO) and endothelin-1 (ET-1). Therefore, this work provides an effective means for the preparation of collagens and novel ACEi peptides from monkfish swim bladders, and the prepared ACEi peptides, including SEGPK, FDGPY and SPGPW, could serve as natural functional components in the development of health care products to control hypertension.

Angiotensin I-Converting Enzyme-Inhibitory Activity and Phytochemical Profile of Constituents of the Leaves of Rehmannia glutinosa f. hueichingensis.

The root of Rehmannia glutinosa Liboschitz forma hueichingensis HSIAO has been used as a tonic and treatment for urinary and skin disorders in Japanese Kampo medicine. Phytochemical investigation of the root has been well reported, but that of the leaves is limited. To explore the potential value of R. glutinosa leaves, we focused on the angiotensin I-converting enzyme (ACE)-inhibitory activity. The leaf extract exhibited ACE-inhibitory activity, and the inhibitory potency of leaves was stronger than that of roots. Using this activity as an indicator, we isolated linaride (1), 6-O-hydroxybenzoyl ajugol (2), acteoside (3), leucosceptoside A (4), martynoside (5), luteolin (6), apigenin (7), and chrysoeriol (8) by separating and purifying the extract. We then examined the ACE-inhibitory activities of 1-8, catalpol (9), aucubin (10), ajugol (11), and echinacoside (12). Among them, 3, 6, and 12 displayed the most potent inhibitory activity. A simultaneous analytical method was also developed using compounds contained in R. glutinosa leaves and roots, and their contents were compared. The method consisted of extraction with 50% aqueous methanol under sonication for 60 min and LC/MS measurement. R. glutinosa leaves tended to have higher levels of majority of the analytes than the roots, including 3 and 6, which had higher ACE-inhibitory activity. These results suggest that 3 and 6 contribute to the ACE-inhibitory activity of R. glutinosa leaves, which may represent a useful medicinal resource for hypertension.

Angiotensin I-Converting Enzyme (ACE) Inhibition and Molecular Docking Study of Meroterpenoids Isolated from Brown Alga, Sargassum macrocarpum.

Angiotensin I-converting enzyme (ACE) is an important blood pressure regulator. In this study, we aimed to investigate the ACE-inhibitory effects of meroterpenoids isolated from the brown alga, Sargassum macrocarpum, and the molecular mechanisms underlying ACE inhibition. Four fractions of S. macrocarpum were prepared using hexane, chloroform, ethyl acetate, and water as solvents and analyzed for their potential ACE-inhibitory effects. The chloroform fraction showed the strongest ACE-inhibitory effect, with an IC50 value of 0.18 mg/mL. Three meroterpenoids, sargachromenol, 7-methyl sargachromenol, and sargaquinoic acid, were isolated from the chloroform fraction. Meroterpenoids isolated from S. macrocarpum had IC50 values of 0.44, 0.37, and 0.14 mM. The molecular docking study revealed that the ACE-inhibitory effect of the isolated meroterpenoids was mainly attributed to Zn-ion, hydrogen bonds, pi-anion, and pi-alkyl interactions between the meroterpenoids and ACE. These results suggest that S. macrocarpum could be a potential raw material for manufacturing antihypertensive nutraceutical ingredients.

In Vitro and In Silico Studies on Angiotensin I-Converting Enzyme Inhibitory Peptides Found in Hydrophobic Domains of Porcine Elastin.

One of the most striking aspects of the primary structure in the hydrophobic domains of the tropoelastin molecule is the occurrence of the VAPGVG repeating sequence. Since the N-terminal tripeptide VAP of VAPGVG showed a potent ACE inhibitory activity, the ACE inhibitory activity of various derivatives of VAP was examined in vitro. The results showed that VAP derivative peptides VLP, VGP, VSP, GAP, LSP, and TRP exhibited potent ACE inhibitory activities, while the non-derivative peptide APG showed only weak activity. In in silico studies, the docking score S value showed that VAP derivative peptides VLP, VGP, VSP, LSP, and TRP had stronger docking interactions than APG. Molecular docking in the ACE active pocket showed that TRP, the most potent ACE inhibitory peptide among the VAP derivatives, had a larger number of interactions with ACE residues in comparison with APG and that the TRP molecule appeared to spread widely in the ACE pocket, while the APG molecule appeared to spread closely. Differences in molecular spread may be a reason why TRP exhibits more potent ACE inhibitory activity than APG. The results suggest that the number and strength of interactions between the peptide and ACE are important for the ACE- inhibitory potency of the peptide.

Functional and Bioactive Properties of Wheat Protein Fractions: Impact of Digestive Enzymes on Antioxidant, α-Amylase, and Angiotensin-Converting Enzyme Inhibition Potential.

This research aimed to determine the biofunctional properties of wheat flour (WF) protein fractions and modifications to the antioxidant, anti-α-amylase and anti-angiotensin-I converting enzyme (ACE) activities induced by the action of digestive endopeptidases in vitro. A molecular characterization of the most abundant protein fractions, i.e., albumins, glutelins-1, glutelins-2 and prolamins, showed that low- and high-MW polypeptides rich in cysteine, glutamic acid and leucine were present in albumins and glutelins, whereas low-MW subunits with a high proportion of polar amino acids prevailed in prolamins. Prolamins exhibited the second-highest water holding capacity (54%) after WF (84%), while albumins provided superior foam stability (76%). Prolamins, glutenins-1 and globulins demonstrated the highest antioxidant activity (up to 95%, 68% and 59%, respectively) both before and after hydrolysis with pepsin (P-H) or trypsin-chymotrypsin (TC-H). Prolamins, globulins and WF strongly inhibited α-amylase (>90%) before and after TC-H, and before P-H (55-71%). Moreover, P-H significantly increased α-amylase inhibition by albumins from 53 to 74%. The fractions with strong ACE inhibitory activity (70-89%) included prolamins and globulins after TC-H or P-H, as well as globulins before TC-H and WF before P-H. This novel evidence indicates that WF protein fractions and their peptide-enriched P and TC hydrolysates are excellent sources of multifunctional bioactives with antioxidant, antihyperglycemic and antihypertensive potential.

Predictive potential of ACE phenotyping in extrapulmonary sarcoidosis.

Elevated ACE expression in tissues (reflected by blood ACE levels) is associated with increased risk of cardiovascular diseases and is also a marker for granulomatous diseases. We developed a new approach for characterization of ACE status in the blood-ACE phenotyping and established normal values of ACE levels 50-150% of control pooled plasma. ACE phenotyping was performed in citrated plasma of 120 patients with known interstitial lung diseases. In the 1st set of 100 patients we found 22 patients with ACE levels > 150%; ACE phenotyping also objectively identified the presence of ACE inhibitors in the plasma of 15 patients. After excluding these patients and patient with ACE mutation that increases ACE shedding, 17 patients were identified as a suspicious for systemic sarcoidosis based on elevation of blood ACE (> 150% of mean). A new parameter that we have established-ACE immunoreactivity (with mAb 9B9)-allowed us to detect 22 patients with decreased values (< 80%) of this parameter, which may indicate the presence of ACE in the blood that originates from macrophages/dendritic cells of granulomas. In the remaining 20 patients, this new parameter (mAbs binding/activity ratio) was calculated using 3 mAbs (9B9, 3A5 and i1A8-having overlapping epitopes), and 8 patients were identified as having decreases in this parameter, thus increasing dramatically the sensitivity for detection of patients with systemic sarcoidosis. Whole body PET scan confirmed extrapulmonary granulomas in some patients with lower immunoreactivity towards anti-ACE mAbs. ACE phenotyping has novel potential to noninvasively detect patients with systemic sarcoidosis.

Angiotensin blockade therapy and survival in pancreatic cancer: a population study.

BACKGROUND: Pancreatic cancer (PC) is one of the most aggressive and challenging cancer types to effectively treat, ranking as the fourth-leading cause of cancer death in the United States. We investigated if exposures to angiotensin II receptor blockers (ARBs) or angiotensin I converting enzyme (ACE) inhibitors after PC diagnosis are associated with survival. METHODS: PC patients were identified by ICD-9 diagnosis and procedure codes among the 3.7 million adults living in the Emilia-Romagna Region from their administrative health care database containing patient data on demographics, hospital discharges, all-cause mortality, and outpatient pharmacy prescriptions. Cox modeling estimated covariate-adjusted mortality hazard ratios for time-dependent ARB and ACE inhibitor exposures after PC diagnosis. RESULTS: 8,158 incident PC patients were identified between 2003 and 2011, among whom 20% had pancreas resection surgery, 36% were diagnosed with metastatic disease, and 7,027 (86%) died by December 2012. Compared to otherwise similar patients, those exposed to ARBs after PC diagnosis experienced 20% lower mortality risk (HR=0.80; 95% CI: 0.72, 0.89). Those exposed to ACE inhibitors during the first three years of survival after PC diagnosis experienced 13% lower mortality risk (HR=0.87; 95% CI: 0.80, 0.94) which attenuated after surviving three years (HR=1.14; 95% CI: 0.90, 1.45). CONCLUSIONS: The results of this large population study suggest that exposures to ARBs and ACE inhibitors after PC diagnosis are significantly associated with improved survival. ARBs and ACE inhibitors could be important considerations for treating PC patients, particularly those with the worst prognosis and most limited treatment options. Considering that these common FDA approved drugs are inexpensive to payers and present minimal increased risk of adverse events to patients, there is an urgent need for randomized clinical trials, large simple randomized trials, or pragmatic clinical trials to formally and broadly evaluate the effects of ARBs and ACE inhibitors on survival in PC patients.

Peptide inhibitors of angiotensin-I converting enzyme based on angiotensin (1-7) with selectivity for the C-terminal domain.

The renin-angiotensin system (RAS) is a key regulator of human arterial pressure. Several of its effects are modulated by angiotensin II, an octapeptide originating from the action of angiotensin-I converting enzyme (ACE) on the decapeptide angiotensin-I. ACE possess two active sites (nACE and cACE) that have their own kinetic and substrate specificities. ACE inhibitors are widely used as the first-line treatment for hypertension and other heart-related diseases, but because they inactivate both ACE domains, their use is associated with serious side effects. Thus, the search for domain-specific ACE inhibitors has been the focus of intense research. Angiotensin (1-7), a peptide that also belongs to the RAS, acts as a substrate of nACE and an inhibitor of cACE. We have synthetized 15 derivatives of Ang (1-7), sequentially removing the N-terminal amino acids and modifying peptides extremities, to find molecules with improved selectivity and inhibition properties. Ac-Ang (2-7)-NH2 is a good ACE inhibitor, resistant to cleavage and with improved cACE selectivity. Molecular dynamics simulations provided a model for this peptide's selectivity, due to Val3 and Tyr4 interactions with ACE subsites. Val3 has an important interaction with the S3 subsite, since its removal greatly reduced peptide-enzyme interactions. Taken together, our findings support ongoing studies using insights from the binding of Ac-Ang (2-7)-NH2 to develop effective cACE inhibitors.

Preparation, Identification, Molecular Docking Study and Protective Function on HUVECs of Novel ACE Inhibitory Peptides from Protein Hydrolysate of Skipjack Tuna Muscle.

To prepare bioactive peptides with high angiotensin-I-converting enzyme (ACE)-inhibitory (ACEi) activity, Alcalase was selected from five kinds of protease for hydrolyzing Skipjack tuna (Katsuwonus pelamis) muscle, and its best hydrolysis conditions were optimized using single factor and response surface experiments. Then, the high ACEi protein hydrolysate (TMPH) of skipjack tuna muscle was prepared using Alcalase under the optimum conditions of enzyme dose 2.3%, enzymolysis temperature 56.2 °C, and pH 9.4, and its ACEi activity reached 72.71% at 1.0 mg/mL. Subsequently, six novel ACEi peptides were prepared from TMPH using ultrafiltration and chromatography methods and were identified as Ser-Pro (SP), Val-Asp-Arg-Tyr-Phe (VDRYF), Val-His-Gly-Val-Val (VHGVV), Tyr-Glu (YE), Phe-Glu-Met (FEM), and Phe-Trp-Arg-Val (FWRV), with molecular weights of 202.3, 698.9, 509.7, 310.4, 425.6, and 606.8 Da, respectively. SP and VDRYF displayed noticeable ACEi activity, with IC50 values of 0.06 ± 0.01 and 0.28 ± 0.03 mg/mL, respectively. Molecular docking analysis illustrated that the high ACEi activity of SP and VDRYF was attributed to effective interaction with the active sites/pockets of ACE by hydrogen bonding, electrostatic force, and hydrophobic interaction. Furthermore, SP and VDRYF could significantly up-regulate nitric oxide (NO) production and down-regulate endothelin-1 (ET-1) secretion in HUVECs after 24 h treatment, but also abolish the negative effect of 0.5 µM norepinephrine (NE) on the generation of NO and ET-1. Therefore, ACEi peptides derived from skipjack tuna (K. pelamis) muscle, especially SP and VDRYF, are beneficial components for functional food against hypertension and cardiovascular diseases.

Screening and Mechanism of Novel Angiotensin-I-Converting Enzyme Inhibitory Peptides in X. sorbifolia Seed Meal: A Computer-Assisted Experimental Study Method.

Angiotensin-I-converting enzyme (ACE) inhibitors are used extensively to control hypertension. In this study, a computer-assisted experimental approach was used to screen ACE-inhibiting peptides from X. sorbifolum seed meal (XSM). The process conditions for XSM hydrolysis were optimized through the orthogonal experimental method combined with a database. The optimal conditions for ACE inhibition included an alkaline protease dose of 5%, 45 °C, 15 min and pH 9.5. The hydrolysate was analyzed by LC-MS/MS, and 10 optimal peptides were screened. Molecular docking results revealed four peptides (GGLPGFDPA, IMAVLAIVL, ETYFIVR, and INPILLPK) with ACE inhibitory potential. At 0.1 mg/mL, the synthetic peptides GGLPGFDPA, ETYFIVR, and INPILLPK provided ACE inhibition rates of 24.89%, 67.02%, and 4.19%, respectively. GGLPGFDPA and ETYFIVR maintained high inhibitory activities during in vitro digestions. Therefore, the XSM protein may be a suitable material for preparing ACE inhibitory peptides, and computer-assisted experimental screening is an effective, accurate and promising method for discovering new active peptides.

Association of the angiotensin I converting enzyme (ACE) gene polymorphisms with recurrent aphthous stomatitis in the Czech population: case-control study.

BACKGROUND: Recurrent aphthous stomatitis (RAS) is multifactorial disease with unclear etiopathogenesis. The aim of this study was to determine distribution of the angiotensin I converting enzyme (ACE) gene polymorphisms and their influence on RAS susceptibility in Czech population. METHODS: The study included 230 subjects (143 healthy controls and 87 patients with RAS) with anamnestic, clinical and laboratory data. Five ACE gene polymorphisms (rs4291/rs4305/rs4311/rs4331/rs1799752 = ACE I/D) were determined by TaqMan technique. RESULTS: The allele and genotype distributions of the studied ACE I/D polymorphisms were not significantly different between subjects with/without RAS (Pcorr > 0.05). However, carriers of II genotype were less frequent in the RAS group (OR = 0.48, 95% CI = 0.21-1.12, P = 0.059). Stratified analysis by sex demonstrated lower frequency of II genotype in women (OR = 0.33, 95% CI = 0.09-1.17, P < 0.035, Pcorr > 0.05, respectively) than in men with RAS (P > 0.05). Moreover, the frequency of AGTGD haplotype was significantly increased in RAS patients (OR = 13.74, 95% CI = 1.70-110.79, P = 0.0012, Pcorr < 0.05). In subanalysis, TGD haplotype was significantly more frequent in RAS patients (P < 0.00001) and CGI haplotype was less frequent in RAS patients (P < 0.01), especially in women (P = 0.016, Pcorr > 0.05). CONCLUSIONS: Our study indicates that while the AGTGD and TGD haplotypes are associated with increased risk of RAS development, CGI haplotype might be one of protective factors against RAS susceptibility in Czech population.

Characterization of Angiotensin I-Converting Enzyme (ACE) inhibitory peptides produced in fermented camel milk (Indian breed) by Lactobacillus acidophilus NCDC-15.

Fermented camel milk provides many health benefits like antidiabetic activity, anti-hypertensive activity etc. Fermented camel milk contains IPP or VPP rich ACE inhibitory peptides. The aim of this study was to spot the novel Angiotensin I-Converting Enzyme inhibitory peptides liberated by the potent proteolytic Lactobacillus acidophilus NCDC-15 from camel milk (Indian breed). NCDC-15 had exhibited maximum PepX activity (0.655) and ACE-inhibitory activity (78.33%) at 12 and 48 h of incubation at 37 °C respectively. Proteolytic activity was measured using o-phthaldialdehyde method and observed maximum (0.976 OD) at 2% of inoculation for 12 h of incubation at 37 °C. Water soluble extracts derived from fermented camel milk were ultrafiltered through 3 kDa, 5 kDa and 10 kDa membrane filters from which 3 kDa permeates (48.01% peptides production & 49.46% ACE-inhibition) and 10 kDa permeates (55.04% peptides production & 42.40% ACE-inhibition) had shown maximum peptides production and ACE-inhibitory activity. Overall, 24 peptides were identified from the samples of 3 kDa permeates [6 fractions (K1, L1, M1, N1, O1 and P1)] and 10 permeates [5 fractions (S, T, U, V and W)]. Novel peptide (AIGPVADLHI) was matched with k-casein in AHTPDB database and other peptides were also found matched with α and ß-caseins of camel milk. Supplementary Information: The online version contains supplementary material available at 10.1007/s13197-022-05357-9.

In vitro and in vivo identification of clinically approved drugs that modify ACE2 expression.

The COVID-19 pandemic caused by SARS-CoV-2 has is a global health challenge. Angiotensin-converting enzyme 2 (ACE2) is the host receptor for SARS-CoV-2 entry. Recent studies have suggested that patients with hypertension and diabetes treated with ACE inhibitors (ACEIs) or angiotensin receptor blockers have a higher risk of COVID-19 infection as these drugs could upregulate ACE2, motivating the study of ACE2 modulation by drugs in current clinical use. Here, we mined published datasets to determine the effects of hundreds of clinically approved drugs on ACE2 expression. We find that ACEIs are enriched for ACE2-upregulating drugs, while antineoplastic agents are enriched for ACE2-downregulating drugs. Vorinostat and isotretinoin are the top ACE2 up/downregulators, respectively, in cell lines. Dexamethasone, a corticosteroid used in treating severe acute respiratory syndrome and COVID-19, significantly upregulates ACE2 both in vitro and in vivo. Further top ACE2 regulators in vivo or in primary cells include erlotinib and bleomycin in the lung and vancomycin, cisplatin, and probenecid in the kidney. Our study provides leads for future work studying ACE2 expression modulators.

SARS-CoV-2 Viral Entry Proteins in Hyperandrogenemic Female Mice: Implications for Women with PCOS and COVID-19.

SARS-CoV-2, the causative agent of COVID-19, infects host cells using the angiotensin I converting enzyme 2 (ACE2) as its receptor after priming by host proteases, including TMPRSS2. COVID-19 affects multiple organ systems, and male patients suffer increased severity and mortality. Polycystic Ovary Syndrome (PCOS) is the most common endocrine disorder in reproductive-age women and is characterized by hyperandrogenism, ovulatory dysfunction, and polycystic ovarian morphology. PCOS is associated with obesity and cardiometabolic comorbidities, both being risk factors associated with severe COVID-19 pathology. We hypothesize that elevated androgens in PCOS regulate SARS-CoV-2 entry proteins in multiple tissues increasing the risk for this population. Female mice were treated with dihydrotestosterone (DHT) for 90 days. Body composition was measured by EchoMRI. Fasting glucose was determined by an enzymatic method. mRNA and protein levels of ACE2, Tmprss2, Cathepsin L, Furin, Tmprss4, and Adam17 were quantified by RT-qPCR, Western-blot, or ELISA in tissues, serum, and urine. DHT treatment increased body weight, fat and lean mass, and fasting glucose. Ace2 mRNA was upregulated in the lung, cecum, heart, and kidney, while downregulated in the brain by DHT. ACE2 protein was upregulated by DHT in the small intestine, heart, and kidney. The SARS-CoV-2 priming proteases Tmprss2, Cathepsin L, and Furin mRNA were upregulated by DHT in the kidney. ACE2 sheddase Adam17 mRNA was upregulated by DHT in the kidney, which corresponded with increased urinary ACE2 in DHT treated mice. Our results highlight the potential for increased cardiac, renal, and gastrointestinal dysfunction in PCOS women with COVID-19.