Role of the left posterior middle temporal gyrus in shape recognition and its reconstruction during drawing: A study combining transcranial magnetic stimulation and functional near infrared spectroscopy.

There are numerous reports of enhanced or emerged visual arts abilities in patients with semantic impairment. These reports led to the theory that a loss of function on the language side of the brain can result in changes of ability to draw and/or to paint. Further, the left posterior middle temporal gyrus (l-pMTG) has been revealed to contribute to the higher control semantic mechanisms with objects recognition and integration of visual information, within a widely distributed network of the left hemisphere. Nevertheless, the theory has not been fully studied in neural bases. The aim of this study is to examine role of the l-pMTG on shape recognition and its reconstruction within drawing behavior, by using a combining method of the repetitive transcranial magnetic stimulation (rTMS) and functional near-infrared spectroscopy (fNIRS). Eighteen healthy participants received a low frequency inhibitory rTMS to their l-pMTG during the drawing task of the Benton Visual Retention Test (BVRT). There was a significant decrease of the mean accuracy of reproductions in the Complex designs of the BVRT, compared to the Simple and Medium designs. The fNIRS data showed strong negative correlations with the results of the BVRT. Though our hypothesis had a contradiction that rTMS would have inhibited the brain activity in the stimulated site, the results suggest that shape recognition and its reconstruction such as the BVRT require neural activations of the l-TL as well as that of the l-pMTG.

Differences in iron balance observed with dietary cholic acid supplementation and marginal iron deficiency in rats.

We investigated whether a cholic acid (CA)-supplemented diet and marginal iron deficiency (MID) diet influence hepatic lipid accumulation and iron balance in rats for 2 weeks. The CA diet enhanced hepatic lipid accumulation and modulated iron metabolism such as enhancement of fecal iron excretion, reduction in iron absorption, and no alteration in plasma iron levels. The MID diet did not alter hepatic lipid concentrations with reduced iron concentration in the liver and plasma. In combination, influence of the CA supplementation on the hepatic iron concentration was opposite between iron-sufficient and MID conditions. In the liver, the CA diet enhanced lipocalin 2 expression, whereas the MID diet enhanced transferrin receptor 1 expression and reduced hepcidin expression. This study revealed an involvement of 12-hydroxylated bile acids in regulation of hepatic iron concentration under MID condition.

A diet supplemented with cholic acid elevates blood pressure accompanied by albuminuria in rats.

A diet supplemented with cholic acid (CA), the primary 12α-hydroxylated bile acid, can induce hepatic lipid accumulation in rats without obesity. This study examined the effects of a CA-supplemented diet on blood pressure (BP). After acclimation, WKAH/HkmSlc rats (3 weeks old) were divided into two groups and fed with a control AIN-93-based diet or a CA-supplemented diet (0.5 g CA/kg) for 13 weeks. The CA diet increased systolic and diastolic BP as well as hepatic lipid concentrations in the rats. No changes were found in the blood sodium concentration. Urinary albumin concentration increased in CA-fed rats. An increase was observed in the hepatic expression of ATP-binding cassette subfamily B member 1B that correlated BPs and urinary albumin concentration accompanied by an increase in portal taurocholic acid concentration. These results suggest that 12α-hydroxylated bile acids are involved in increased BP and albuminuria via alteration of hepatic function.

The ratio of 12α to non-12-hydroxylated bile acids reflects hepatic triacylglycerol accumulation in high-fat diet-fed C57BL/6J mice.

In our previous study, enterohepatic 12α-hydroxylated (12α) bile acid (BA) levels were found to be correlated with hepatic triacylglycerol concentration in rats fed high-fat (HF) diet. Since BA composition is diverse depending on animal species, we evaluated whether such a relationship is applicable in mice in response to an HF diet. C57BL/6JJmsSLC (B6) male mice were fed HF diet for 13 weeks and analyzed for triacylglycerol, cholesterol, oxysterols, and other metabolites in the liver. The BA composition was determined in the liver, small intestinal contents, portal plasma, aortic plasma, and feces. Neutral sterols were also measured in the feces. The ratio of 12α BA/non-12 BA increased in the liver, portal plasma, small intestinal contents, and feces of HF-fed B6 mice. Moreover, a positive correlation was observed between the ratio of fecal 12α BAs/non-12 BAs and hepatic triacylglycerol concentration. The concentration of 7α-hydroxycholesterol was increased in the liver of HF-fed B6 mice, whereas no increase was observed in the hepatic expression of cytochrome P450 family 7 subfamily A member 1. The present study showed that the ratio of 12α BA/non-12 BA in feces is closely associated with hepatic triacylglycerol accumulation in B6 mice fed HF diet.

Taurocholic acid, a primary 12α-hydroxylated bile acid, induces leakiness in the distal small intestine in rats.

A high-fat diet increases 12α-hydroxylated (12αOH) bile acid (BA) secretion in rats, and secondary BAs are responsible for the leaky gut. This study aimed to examine the role of primary 12αOH BAs in gut barrier impairment in rats using dietary cholic acid (CA) supplementation (0.5 g/kg diet). The CA diet increased the 12αOH BAs concentrations in the small and large intestine, accompanied by gut barrier impairment. Based on the luminal 12αOH BAs concentrations, ex vivo gut leakiness was determined. Deoxycholic acid increased permeability in the large intestine, whereas taurocholic acid (TCA) increased the ileal permeability, but not jejunal permeability. A Rho kinase inhibitor attenuated TCA-induced ileal permeability. Administration of vancomycin, which abolishes secondary BAs, did not influence the gut leakiness induced by the CA diet. Changes in the gut permeation marker in the tail vein blood suggested the possibility that the CA-induced leakiness occurred in the small intestine. The CA diet enhanced the phosphorylation of myosin light chain 2 and reduced claudins expressions in rat ileal epithelia. Reductions in barrier function-related genes were observed in the ileum, but not in the colon of the CA-fed rats. Overall, the present study demonstrated the significance of TCA in proximal gut leakiness.

12α-Hydroxylated bile acid enhances accumulation of adiponectin and immunoglobulin A in the rat ileum.

We previously reported that dietary supplementation with cholic acid (CA), the primary 12α-hydroxylated (12αOH) bile acid (BA), reduces plasma adiponectin concentration in rats. The aim of this study was to examine the distribution of adiponectin in the body of CA-fed rats and its influence on mucosal immunoglobulin A concentration in the intestine. Rats were fed a diet supplemented with or without CA (0.5 g CA/kg diet) for 13 weeks. A reduction in plasma adiponectin level was observed from week 3. At the end of the experiment, the CA diet reduced plasma adiponectin concentration both in the portal and aortic plasma. Accumulation of adiponectin was accompanied by an increase in cadherin-13 mRNA expression in the ileal mucosa of CA-fed rats. No increase was observed in adiponectin mRNA expression in the ileal and adipose tissues of the CA-fed rats. Immunoglobulin A concentration in the ileal mucosa was elevated in the CA-fed rats and was correlated with the ileal adiponectin concentration. 12αOH BAs may modulate mucosal immune response that are involved in the accumulation of adiponectin in the ileum.

Primary 12α-Hydroxylated Bile Acids Lower Hepatic Iron Concentration in Rats.

BACKGROUND: Primary 12α-hydroxylated bile acids (12αOH BAs) enhance intestinal iron uptake due to their ability ex vivo to chelate iron. However, no information is available on their role in vivo, especially in the liver. OBJECTIVES: To investigate the effects and mechanisms of primary 12αOH BAs on hepatic iron concentration in vivo. METHODS: Male Wistar King A Hokkaido male rats (WKAH/HkmSlc) rats aged 4-5 weeks were fed a control diet or a diet with cholic acid (CA; 0.5 g/kg diet), the primary 12αOH BA, for 2 weeks (Study 1) or 13 weeks (Study 2). In Study 3, rats fed the same diets were given drinking water either alone or containing vancomycin (200 mg/L) for 6 weeks. The variables measured included food intake (Studies 1-3), bile acid profiles (Studies 1 and 3), hepatic iron concentration (Studies 1-3), fecal iron excretion (Studies 1 and 2), iron-related liver gene expression (Studies 2 and 3), and plasma iron-related factors (Studies 2 and 3). RESULTS: In Study 1, CA feed reduced the hepatic iron concentration (-16%; P = 0.005) without changing food intake or fecal iron excretion. In Study 2, we found a significant increase in the aortic plasma concentration of lipocalin 2 (LCN2; +65%; P < 0.001), an iron-trafficking protein. In Study 3, we observed no effect of vancomycin treatment on the CA-induced reduction of hepatic iron concentration (-32%; P < 0.001), accompanied by increased plasma LCN2 concentration (+72%; P = 0.003), in the CA-fed rats despite a drastic reduction in the secondary 12αOH BA concentration (-94%; P < 0.001) in the aortic plasma. CONCLUSIONS: Primary 12αOH BAs reduced the hepatic iron concentration in rats. LCN2 may be responsible for the hepatic iron-lowering effect of primary 12αOH BAs by transporting iron out of the liver.

12α-Hydroxylated bile acid induces hepatic steatosis with dysbiosis in rats.

There is an increasing need to explore the mechanism of the progression of non-alcoholic fatty liver disease. Steroid metabolism is closely linked to hepatic steatosis and steroids are excreted as bile acids (BAs). Here, we demonstrated that feeding WKAH/HkmSlc inbred rats a diet supplemented with cholic acid (CA) at 0.5 g/kg for 13 weeks induced simple steatosis without obesity. Liver triglyceride and cholesterol levels were increased accompanied by mild elevation of aminotransferase activities. There were no signs of inflammation, insulin resistance, oxidative stress, or fibrosis. CA supplementation increased levels of CA and taurocholic acid (TCA) in enterohepatic circulation and deoxycholic acid (DCA) levels in cecum with an increased ratio of 12α-hydroxylated BAs to non-12α-hydroxylated BAs. Analyses of hepatic gene expression revealed no apparent feedback control of BA and cholesterol biosynthesis. CA feeding induced dysbiosis in cecal microbiota with enrichment of DCA producers, which underlines the increased cecal DCA levels. The mechanism of steatosis was increased expression of Srebp1 (positive regulator of liver lipogenesis) through activation of the liver X receptor by increased oxysterols in the CA-fed rats, especially 4ß-hydroxycholesterol (4ßOH) formed by upregulated expression of hepatic Cyp3a2, responsible for 4ßOH formation. Multiple regression analyses identified portal TCA and cecal DCA as positive predictors for liver 4ßOH levels. The possible mechanisms linking these predictors and upregulated expression of Cyp3a2 are discussed. Overall, our observations highlight the role of 12α-hydroxylated BAs in triggering liver lipogenesis and allow us to explore the mechanisms of hepatic steatosis onset, focusing on cholesterol and BA metabolism.

A low coefficient of variation in hepatic triglyceride concentration in an inbred rat strain.

BACKGROUND: Inbred strains are characterized by less genetic variation, which suggests usefulness of inbred strains for evaluations of various parameters. In this study, experimental reproducibility in several parameters was compared between an outbred Wistar rat and Wistar King A Hokkaido (WKAH/HkmSlc) rat, the inbred strain that is originated from Wistar rats. METHODS: Difference of variations was investigated in parameters of body compositions and liver functions such as body weight, liver weight, liver triglycerides (TG), liver cholesterol and plasma alanine aminotransferase activity (ALT) between WKAH rats and outbred Wistar rats by using the coefficient of variation (CV). RESULTS: There was no difference in the CVs of body weight and relative liver weight between WKAH and Wistar rats. The CVs of body weight and relative liver weight were below 10% in both WKAH and Wistar rats. The CVs of TG, cholesterol, and ALT in Wistar rats were between 30 and 40%, whereas those in WKAH rats were between 10 and 25%. A low CV level of TG was observed in WKAH rats compared to that in Wistar rats regardless of the duration of the experimental period in those rat strains. CONCLUSION: The low CV values in metabolic parameters involved in liver functions in the inbred rats suggested an advantage of using inbred rather than outbred rats for the evaluation of liver lipid metabolism.

Association between 12α-hydroxylated bile acids and hepatic steatosis in rats fed a high-fat diet.

High-fat (HF) diet induces hepatic steatosis that is a risk factor for noncommunicable diseases such as obesity, type 2 diabetes and cardiovascular disease. Previously, we found that HF feeding in rats increases the excretion of fecal bile acids (BAs), specifically 12α-hydroxylated (12αOH) BAs. Although the liver is the metabolic center in our body, the association between hepatic steatosis and 12αOH BAs in HF-fed rats is unclear. Thus, we investigated extensively BA composition in HF-fed rats and evaluated the association between hepatic steatosis and 12αOH BAs. Acclimated male inbred WKAH/HkmSlc rats were divided into two groups and fed either control or HF diet for 8 weeks. Feeding HF diet increased hepatic triglyceride and total cholesterol concentrations, which correlated positively with 12αOH BAs concentrations but not with non-12αOH BAs in the feces, portal plasma and liver. Accompanied by the increase in 12αOH BAs, the rats fed HF diet showed increased fat absorption and higher mRNA expression of liver Cidea. The enhancement of 12αOH BA secretion may contribute to hepatic steatosis by the promotion of dietary fat absorption and hepatic Cidea mRNA expression. The increase in 12αOH BAs was associated with enhanced liver cholesterol 7α-hydroxylase (Cyp7a1) and sterol 12α-hydroxylase (Cyp8b1) mRNA expression. There was a significant increase in 7α-hydroxycholesterol, a precursor of BAs, in the liver of HF-fed rats. Altogether, these data suggest that the HF diet increases preferentially 12αOH BAs synthesis by utilizing the accumulated hepatic cholesterol and enhancing mRNA expression of Cyp7a1 and Cyp8b1 in the liver.

Lithocholic acid increases intestinal phosphate and calcium absorption in a vitamin D receptor dependent but transcellular pathway independent manner.

Phosphate/calcium homeostasis is crucial for health maintenance. Lithocholic acid, a bile acid produced by intestinal bacteria, is an agonist of vitamin D receptor. However, its effects on phosphate/calcium homeostasis remain unclear. Here, we demonstrated that lithocholic acid increases intestinal phosphate/calcium absorption in an enterocyte vitamin D receptor-dependent manner. Lithocholic acid was found to increase serum phosphate/calcium levels and thus to exacerbate vascular calcification in animals with chronic kidney disease. Lithocholic acid did not affect levels of intestinal sodium-dependent phosphate transport protein 2b, Pi transporter-1, -2, or transient receptor potential vanilloid subfamily member 6. Everted gut sac analyses demonstrated that lithocholic acid increased phosphate/calcium absorption in a transcellular pathway-independent manner. Lithocholic acid suppressed intestinal mucosal claudin 3 and occludin in wild-type mice, but not in vitamin D receptor knockout mice. Everted gut sacs of claudin 3 knockout mice showed an increased permeability for phosphate, but not calcium. In patients with chronic kidney disease, serum 1,25(OH)2 vitamin D levels are decreased, probably as an intrinsic adjustment to reduce phosphate/calcium burden. In contrast, serum and fecal lithocholic acid levels and fecal levels of bile acid 7α-dehydratase, a rate-limiting enzyme involved in lithocholic acid production, were not downregulated. The effects of lithocholic acid were eliminated by bile acid adsorptive resin in mice. Thus, lithocholic acid and claudin 3 may represent novel therapeutic targets for reducing phosphate burden.

Untargeted Lipidomic Analysis of Plasma from High-fat Diet-induced Obese Rats Using UHPLC-Linear Trap Quadrupole-Orbitrap MS.

High-fat diet (HFD)-induced obesity is a primary risk factor for serious health problems. Although much research has been performed at the genomic level, lipidomic studies were limited. In this study, we aim to obtain a comprehensive profile of circulating plasma lipids, which are altered in rodent rat obesity by untargeted liquid chromatography-mass spectrometry. Rats fed with HFD for 8 weeks had increased body weight, liver and adipose tissue weight. The analysis results revealed that polyunsaturated fatty acids (PUFAs) and their corresponding phosphatidylcholine, phosphatidylinositol, and phosphatidylserine were significantly decreased in rats fed with HFD. In contrast, less unsaturated and ether type phosphatidylglycerols were increased. The triacylglycerides (TAGs) having saturated FA were increased in the HFD condition, whereas TAGs having PUFA were decreased. The levels of many plasma lipids were altered, and interestingly PUFA derived lipids were negatively associated with obesity. This signifies the importance of a PUFAs enriched diet to overwhelm obesity associated diseases.

Correlation between 12α-hydroxylated bile acids and insulin secretion during glucose tolerance tests in rats fed a high-fat and high-sucrose diet.

BACKGROUND: Previously, we found a significant relationship in a rat study between energy intake and bile acid (BA) metabolism especially 12α-hydroxylated (12αOH) BAs. The present study was designed to reveal relationships among BA metabolism, glucose tolerance, and cecal organic acids in rats fed a high-fat and high-sucrose diet (HFS) by using multivariate and multiple regression analyses in two types of glucose tolerance tests (GTTs). METHODS: Male WKAH/HkmSlc rats were fed with a control or a HFS for 13 weeks. Oral glucose tolerance test (OGTT) and intraperitoneal glucose tolerance test (IPGTT) were performed at week 9 and 11, respectively. BAs were analyzed by using ultra high-performance liquid chromatography-mass spectrometry. Organic acid concentrations in cecal contents were analyzed by using ultra high-performance liquid chromatography with post-column pH buffered electric conductivity method. RESULTS: A positive correlation of aortic 12αOH BA concentration was observed with energy intake and visceral adipose tissue weight. We found that an increase of 12αOH BAs in enterohepatic circulation, intestinal contents and feces in the HFS-fed rats compared to those in control rats regardless of no significant increase of total BA concentration in the feces in the test period. Fecal 12αOH BA concentration was positively correlated with maximal insulin level in OGTT and area under curve of insulin in IPGTT. There was a positive correlation between aortic 12αOH BAs concentration and changes in plasma glucose level in both OGTT and IPGTT. In contrast, a decrease in the concentration of organic acids was observed in the cecal contents of the HFS-fed rats. Multiple linear regression analysis in the IPGTT revealed that the concentrations of aortic 12αOH BA and cecal acetic acid were the predictors of insulin secretion. Moreover, there was a positive correlation between concentration of portal 12αOH BAs and change in insulin concentration of peripheral blood in the IPGTT. CONCLUSION: The distribution analysis of BA compositions accompanied by GTTs revealed a close relationship between 12αOH BA metabolism and insulin secretion in GTTs in rats.

Ingestion of difructose anhydride III partially suppresses the deconjugation and 7α-dehydroxylation of bile acids in rats fed with a cholic acid-supplemented diet.

Difructose anhydride III (DFAIII) is a prebiotic involved in the reduction of secondary bile acids (BAs). We investigated whether DFAIII modulates BA metabolism, including enterohepatic circulation, in the rats fed with a diet supplemented with cholic acid (CA), one of the 12α-hydroxylated BAs. After acclimation, the rats were fed with a control diet or a diet supplemented with DFAIII. After 2 weeks, each group was further divided into two groups and was fed diet with or without CA supplementation at 0.5 g/kg diet. BA levels were analyzed in aortic and portal plasma, liver, intestinal content, and feces. As a result, DFAIII ingestion reduced the fecal deoxycholic acid level via the partial suppression of deconjugation and 7α-dehydroxylation of BAs following CA supplementation. These results suggest that DFAIII suppresses production of deoxycholic acid in conditions of high concentrations of 12α-hydroxylated BAs in enterohepatic circulation, such as obesity or excess energy intake. Abbreviation: BA: bile acid; BSH: bile salt hydrolase; CA: cholic acid; DCA: deoxycholic acid; DFAIII: difructose anhydride III; MCA: muricholic acid; MS: mass spectrometry; NCDs: non-communicable diseases; LC: liquid chromatography; SCFA: short-chain fatty acid; TCA: taurocholic acid; TCDCA: taurochenodeoxycholic acid; TDCA: taurodeoxycholic acid; TUDCA: tauroursodeoxychlic acid; TαMCA: tauro-α-muricholic acid; TßMCA: tauro-ß-muricholic acid; TωMCA: tauro-ω-muricholic acid.

Alteration of Bile Acid Metabolism by a High-Fat Diet Is Associated with Plasma Transaminase Activities and Glucose Intolerance in Rats.

Ingestion of a high-fat (HF) diet is known to enhance bile acid (BA) secretion, but precise information about the BA molecular species is lacking, especially information on the conjugated BAs in enterohepatic circulation. As cholesterol is the precursor of BAs, we analyzed alterations of the entire BA metabolic pathway in response to a HF diet without the addition of cholesterol and BA in the diet. Additionally, we evaluated the relationships between BA metabolism and some disorders, such as plasma transaminase activities and glucose intolerance induced by the HF diet. Acclimated WKAH/HkmSlc male rats (3 wk old) were divided into two groups fed a control or the HF diet for 22 wk. Fasting blood glucose was measured during the experimental period, and an intraperitoneal glucose tolerance test was performed at week 21. As a result, ingestion of the HF diet selectively increased the concentration of taurocholic acid in the bile and small intestinal contents as well as deoxycholic acid in the large intestinal contents and feces. These results indicated a selective increase of 12α-hydroxylated BA concentrations in response to the HF diet. Moreover, fecal 12α-hydroxylated BA concentration was positively correlated with cumulative energy intake, visceral adipose tissue weight, and glucose intolerance. The present study suggests that fecal 12α-hydroxylated BA is a non-invasive marker that can detect the early phase of glucose intolerance.

Marginal iron deficiency enhances liver triglyceride accumulation in rats fed a high-sucrose diet.

We investigated whether marginal iron-deficiency (MID) without anemia influences liver lipid accumulation in rats. Ingestion of a MID diet in which the iron concentration was half of AIN-93 formulation (iron-adequate, IA) for 3 weeks decreased liver iron concentration without anemia. We then evaluated the influence of the MID diet on liver lipid accumulation in combination with a high-sucrose (HS) diet and confirmed that the HS-MID diet successfully decreased liver iron concentration without anemia. Additionally, a significant increase in liver triglyceride concentration was found, accompanied by upregulation of hepatic fatty acid synthase expression in the rats fed the HS-MID diet compared to those in the rats fed an HS-IA diet, although no difference was observed in plasma transaminase activity and hepatic interleukin-1ß expression. These results suggest that MID enhances de novo lipid synthesis via upregulation of lipogenic gene expression in combination with sucrose in the diet. Abbreviations: ALT, alanine aminotransferase; AST, aspartate aminotransferase; HS, high sucrose; IA, iron adequate; ID, iron deficiency; MID, marginal irondeficiency; NAFLD, non-alcoholic fatty liver disease.

Effects of Light and Sound on the Prefrontal Cortex Activation and Emotional Function: A Functional Near-Infrared Spectroscopy Study.

We constructed a near infrared spectroscopy-based real-time feedback system to estimate the subjects' emotional states using the changes in oxygenated hemoglobin concentration [Δ(oxy-Hb)] in the prefrontal cortex (PFC). Using this system, we investigated the influences of continual mild and equivocal stimuli consisting of lights and a reconstructed waterfall sound on Δ[oxy-Hb] in the PFC. The visual (light) and auditory (sound) stimuli changed randomly and independently, depending on the emotional states of the individual subjects. The emotional states induced by the stimuli were examined via a questionnaire rated on an 11-point scale, from +5 (pleasant) to -5 (unpleasant), through 0 (neutral), after the 5-min experiments. Results from 757 subjects revealed that Δ[oxy-Hb] in the PFC exhibited a weak, but significant, correlation with emotional change, with the given continual and mild stimuli similar to that experienced in response to the intense pleasant/unpleasant stimuli. Based on the results we discuss the generation of pleasant/unpleasant weak emotional change induced by mild and weak stimuli such as light and sound.

Influence of subjective happiness on the prefrontal brain activity: an fNIRS study.

Focusing on the relationship between subjective happiness (SH) and emotional changes, we examined influences of SH on emotion-related prefrontal activity using multichannel NIRS. The International Affective Picture System (IAPS) was used to evoke emotional changes. Subjects were a total of 18 right-handed healthy students. Frequency of picture-induced increases in oxygenated haemoglobin (oxy-Hb) was evaluated. Subjects with a high SH score had a higher frequency of increased oxy-Hb in the left prefrontal cortex (PFC) while viewing pleasant pictures, whereas they showed a lower frequency in the right PFC while viewing unpleasant pictures. It is well known that the left PFC and right PFC are engaged in different ways in the emotional processes. Although further investigations are required, the present results indicate that the SH level influences the right-left differences in emotion-related prefrontal activity.

Two arginine residues in the substrate pocket predominantly control the substrate selectivity of thiocyanate hydrolase.

Thiocyanate hydrolase (SCNase) of Thiobacillus thioparus THI115 is a cobalt (Co)-containing enzyme that catalyzes the hydrolysis of thiocyanate (SCN⁻), a major component of wastewater from coke oven factories, to carbonyl sulfide and ammonia. Although SCNase exhibits high structural similarities to Co-type nitrile hydratase (NHase), including a unique Co³âº catalytic center with two oxidized Cys ligands, both SCNase and NHase exclusively catalyze only their own substrates. Based on the differences in the substrate-binding pockets of these enzymes, ßArg90 and γArg136 of SCNase, with side chains extending toward the pocket, were separately substituted with Phe and Trp, the corresponding residues, respectively, in Co-type NHase. Both SCNase ßArg90 and SCNase γArg136 mutants showed no SCN⁻ hydrolysis activity but did catalyze the hydration of nitriles. The estimated kcat values (∼2 s⁻¹) corresponded to approximately 0.2% of that of Co-type NHase for nitrile hydration and approximately 3% of that of wild-type SCNase for SCN⁻ hydrolysis. The crystal structure of SCNase γR136W is essentially identical to that of the wild-type, including the Co³âº center having Cys oxidations; the size of the substrate pocket was enlarged because of conformational changes on the side chains of the mutated residue. Discussion of the difference in the environments around the substrate-binding pockets among the wild-type and mutant SCNases and Co-type NHase strongly suggests that ßArg90 and γArg136, positioned at the top of the Co³âº center, predominantly control the substrate selectivity of SCNase.