Regulating bile acids signaling for NAFLD: molecular insights and novel therapeutic interventions.

Nonalcoholic fatty liver disease (NAFLD) emerges as the most predominant cause of liver disease, tightly linked to metabolic dysfunction. Bile acids (BAs), initially synthesized from cholesterol in the liver, undergo further metabolism by gut bacteria. Increasingly acknowledged as critical modulators of metabolic processes, BAs have been implicated as important signaling molecules. In this review, we will focus on the mechanism of BAs signaling involved in glucose homeostasis, lipid metabolism, energy expenditure, and immune regulation and summarize their roles in the pathogenesis of NAFLD. Furthermore, gut microbiota dysbiosis plays a key role in the development of NAFLD, and the interactions between BAs and intestinal microbiota is elucidated. In addition, we also discuss potential therapeutic strategies for NAFLD, including drugs targeting BA receptors, modulation of intestinal microbiota, and metabolic surgery.

Targeting the gut microbiota and its metabolites for type 2 diabetes mellitus.

Type 2 diabetes mellitus (T2DM) is a metabolic disorder characterized by hyperglycemia and insulin resistance. The incidence of T2DM is increasing globally, and a growing body of evidence suggests that gut microbiota dysbiosis may contribute to the development of this disease. Gut microbiota-derived metabolites, including bile acids, lipopolysaccharide, trimethylamine-N-oxide, tryptophan and indole derivatives, and short-chain fatty acids, have been shown to be involved in the pathogenesis of T2DM, playing a key role in the host-microbe crosstalk. This review aims to summarize the molecular links between gut microbiota-derived metabolites and the pathogenesis of T2DM. Additionally, we review the potential therapy and treatments for T2DM using probiotics, prebiotics, fecal microbiota transplantation and other methods to modulate gut microbiota and its metabolites. Clinical trials investigating the role of gut microbiota and its metabolites have been critically discussed. This review highlights that targeting the gut microbiota and its metabolites could be a potential therapeutic strategy for the prevention and treatment of T2DM.

Sustainable and efficient method utilizing N-acetyl-L-cysteine for complete and enhanced ochratoxin A clearance by antagonistic yeast.

With the increasing global climate change, ochratoxin A (OTA) pollution in food and environment has become a serious and potential risk element threatening food safety and human health. Biodegradation of mycotoxin is an eco-friendly and efficient control strategy. Still, research works are warranted to develop low-cost, efficient, and sustainable approaches to enhance the mycotoxin degradation efficiency of microorganisms. In this study, the activities of N-acetyl-L-cysteine (NAC) against OTA toxicity were evidenced, and its positive effects on the OTA degradation efficiency of antagonistic yeast, Cryptococcus podzolicus Y3 were verified. Co-culturing C. podzolicus Y3 with 10 mM NAC improved 100% and 92.6% OTA degradation rate into ochratoxin α (OTα) at 1 d and 2 d. The excellent promotion role of NAC on OTA degradation was observed even at low temperatures and alkaline conditions. C. podzolicus Y3 treated with OTA or OTA+NAC promoted reduced glutathione (GSH) accumulation. GSS and GSR genes were highly expressed after OTA and OTA+NAC treatment, contributing to GSH accumulation. In the early stages of NAC treatment, yeast viability and cell membrane were reduced, but the antioxidant property of NAC prevented lipid peroxidation. Our finding provides a sustainable and efficient new strategy to improve mycotoxin degradation by antagonistic yeasts, which could be applied to mycotoxin clearance.

[Corrigendum] High glucose and high insulin conditions promote MCF­7 cell proliferation and invasion by upregulating IRS1 and activating the Ras/Raf/ERK pathway.

Subsequently to the publication of this paper, an interested reader drew to the authors' attention that, in the wound­healing assays portrayed in Fig. 2A on p. 6692, in the 0 h row, the 'NG + LI' and 'HG + HI' panels contained overlapping data, such that they appeared to have been derived from the same original source. After having examined their original data, the authors have realized that this figure was inadvertently assembled incorrectly. The corrected version of Fig. 2. showing the correct data for the 'HG + HI' panel, is shown on the next page. Note that this error did not significantly affect the results or the conclusions reported in this paper, and all the authors agree with the publication of this Corrigendum. Furthermore, the authors apologize to the readership for any inconvenience caused. [Molecular Medicine Reports 16: 6690­6696, 2017; DOI: 10.3892/mmr.2017.7420].

Identification of Key Biomarkers in Systemic Lupus Erythematosus by a Multi-Cohort Analysis.

Systemic lupus erythematosus (SLE) is an autoimmune disease that affects multiple body systems with heterogeneous clinical manifestations. Since gene expression analyses have been accomplished on diverse types of samples to specify SLE-related genes, single-cohort transcriptomics have not produced reliable results. Using an integrated multi-cohort analysis framework, we analyzed whole blood cells from SLE patients from three transcriptomics cohorts (n=1222) and identified a five-gene signature that distinguished SLE patients from controls. We validated the diagnostic performance of this five-gene signature in six independent validation cohorts (n= 469), with an area under the receiver operating characteristic curve of 0.88 [95% CI 0.7 - 0.96]. This five-gene signature may be associated with the proportion of SLE immune cells, and generalizable across ages and sample types with real diagnostic value for clinical application.

Association of Septic Shock with Mortality in Hospitalized COVID-19 Patients in Wuhan, China.

Purpose: Septic shock is a severe complication of COVID-19 patients. We aim to identify risk factors associated with septic shock and mortality among COVID-19 patients. Methods: A total of 212 COVID-19 confirmed patients in Wuhan were included in this retrospective study. Clinical outcomes were designated as nonseptic shock and septic shock. Log-rank test was conducted to determine any association with clinical progression. A prediction model was established using random forest. Results: The mortality of septic shock and nonshock patients with COVID-19 was 96.7% (29/30) and 3.8% (7/182). Patients taking hypnotics had a much lower chance to develop septic shock (HR = 0.096, p=0.0014). By univariate logistic regression analysis, 40 risk factors were significantly associated with septic shock. Based on multiple regression analysis, eight risk factors were shown to be independent risk factors and these factors were then selected to build a model to predict septic shock with AUC = 0.956. These eight factors included disease severity (HR = 15, p < 0.001), age > 65 years (HR = 2.6, p=0.012), temperature > 39.1°C (HR = 2.9, p=0.047), white blood cell count > 10 × 109 (HR = 6.9, p < 0.001), neutrophil count > 75 × 109 (HR = 2.4, p=0.022), creatine kinase > 5 U/L (HR = 1.8, p=0.042), glucose > 6.1 mmol/L (HR = 7, p < 0.001), and lactate > 2 mmol/L (HR = 22, p < 0.001). Conclusions: We found 40 risk factors were significantly associated with septic shock. The model contained eight independent factors that can accurately predict septic shock. The administration of hypnotics could potentially reduce the incidence of septic shock in COVID-19 patients.

Gut microbiota-bile acid crosstalk contributes to the rebound weight gain after calorie restriction in mice.

Calorie restriction (CR) and fasting are common approaches to weight reduction, but the maintenance is difficult after resuming food consumption. Meanwhile, the gut microbiome associated with energy harvest alters dramatically in response to nutrient deprivation. Here, we reported that CR and high-fat diet (HFD) both remodeled the gut microbiota with similar microbial composition, Parabacteroides distasonis was most significantly decreased after CR or HFD. CR altered microbiota and reprogramed metabolism, resulting in a distinct serum bile acid profile characterized by depleting the proportion of non-12α-hydroxylated bile acids, ursodeoxycholic acid and lithocholic acid. Downregulation of UCP1 expression in brown adipose tissue and decreased serum GLP-1 were observed in the weight-rebound mice. Moreover, treatment with Parabacteroides distasonis or non-12α-hydroxylated bile acids ameliorated weight regain via increased thermogenesis. Our results highlighted the gut microbiota-bile acid crosstalk in rebound weight gain and Parabacteroides distasonis as a potential probiotic to prevent rapid post-CR weight gain.

Hydrogen sulfide restores sevoflurane postconditioning mediated cardioprotection in diabetic rats: Role of SIRT1/Nrf2 signaling-modulated mitochondrial dysfunction and oxidative stress.

Diabetic hearts are vulnerable to myocardial ischemia/reperfusion injury (IRI), but are insensitive to sevoflurane postconditioning (SPC), activating peroxiredoxins that confer cardioprotection. Previous studies have demonstrated that hydrogen sulfide (H2 S) can suppress oxidative stress of diabetic rats through increasing the expression of silent information regulator factor 2-related enzyme 1 (SIRT1), but whether cardioprotection by SPC can be restored afterward remains unclear. Diabetic rat was subjected to IRI (30 min of ischemia followed by 120 min reperfusion). Postconditioning treatment with sevoflurane was administered for 15 min upon the onset of reperfusion. The diabetic rats were treated with GYY4137 (H2 S donor) 5 days before the experiment. Myocardial infarct size, mitochondrial structure and function, ATP content, activities of complex I-IV, marker of oxidative stress, SIRT1, nuclear factor E2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and NADPH Oxidase-2 (Nox-2) protein expression were detected after reperfusion, and cardiac function was evaluated by echocardiography at 24 h after reperfusion. After H2 S activated SIRT1 in the impaired myocardium of diabetic rats, SPC significantly upregulated the expression of Nrf2 and its downstream mediator HO-1, thus reduced the expression of Nox-2. In addition, H2 S remarkably increased cytoplasmic and nuclear SIRT1 which was further enhanced by SPC. Furthermore, H2 S combined with SPC reduced the production of reactive oxygen species, increased the content of ATP, and maintained mitochondrial enzyme activity. Finally, myocardial infarct size and myocardium damage were decreased, and cardiac function was improved. Taken together, our study proved that H2 S could restore SPC-induced cardioprotection in diabetic rats by enhancing and promoting SIRT1/Nrf2 signaling pathway mediated mitochondrial dysfunction and oxidative stress.

Targeting the alternative bile acid synthetic pathway for metabolic diseases.

The gut microbiota is profoundly involved in glucose and lipid metabolism, in part by regulating bile acid (BA) metabolism and affecting multiple BA-receptor signaling pathways. BAs are synthesized in the liver by multi-step reactions catalyzed via two distinct routes, the classical pathway (producing the 12α-hydroxylated primary BA, cholic acid), and the alternative pathway (producing the non-12α-hydroxylated primary BA, chenodeoxycholic acid). BA synthesis and excretion is a major pathway of cholesterol and lipid catabolism, and thus, is implicated in a variety of metabolic diseases including obesity, insulin resistance, and nonalcoholic fatty liver disease. Additionally, both oxysterols and BAs function as signaling molecules that activate multiple nuclear and membrane receptor-mediated signaling pathways in various tissues, regulating glucose, lipid homeostasis, inflammation, and energy expenditure. Modulating BA synthesis and composition to regulate BA signaling is an interesting and novel direction for developing therapies for metabolic disease. In this review, we summarize the most recent findings on the role of BA synthetic pathways, with a focus on the role of the alternative pathway, which has been under-investigated, in treating hyperglycemia and fatty liver disease. We also discuss future perspectives to develop promising pharmacological strategies targeting the alternative BA synthetic pathway for the treatment of metabolic diseases.

A dysregulated bile acid-gut microbiota axis contributes to obesity susceptibility.

BACKGROUND: The composition of the bile acid (BA) pool is closely associated with obesity and is modified by gut microbiota. Perturbations of gut microbiota shape the BA composition, which, in turn, may alter important BA signaling and affect host metabolism. METHODS: We investigated BA composition of high BMI subjects from a human cohort study and a high fat diet (HFD) obesity prone (HF-OP) / HFD obesity resistant (HF-OR) mice model. Gut microbiota was analysed by metagenomics sequencing. GLP-1 secretion and gene regulation studies involved ELISA, qPCR, Western blot, Immunohistochemistry, and Immunofluorescence staining. FINDINGS: We found that the proportion of non-12-OH BAs was significantly decreased in the unhealthy high BMI subjects. The HF-OR mice had an enhanced level of non-12-OH BAs. Non-12-OH BAs including ursodeoxycholate (UDCA), chenodeoxycholate (CDCA), and lithocholate (LCA) were decreased in the HF-OP mice and associated with altered gut microbiota. Clostridium scindens was decreased in HF-OP mice and had a positive correlation with UDCA and LCA. Gavage of Clostridium scindens in mice increased the levels of hepatic non-12-OH BAs, accompanied by elevated serum 7α-hydroxy-4-cholesten-3-one (C4) levels. In HF-OP mice, altered BA composition was associated with significantly downregulated expression of GLP-1 in ileum and PGC1α, UCP1 in brown adipose tissue. In addition, we identified that UDCA attenuated the high fat diet-induced obesity via enhancing levels of non-12-OH BAs. INTERPRETATION: Our study highlights that dysregulated BA signaling mediated by gut microbiota contributes to obesity susceptibility, suggesting modulation of BAs could be a promising strategy for obesity therapy.

Postharvest acibenzolar-S-methyl treatment maintains storage quality and retards softening of apple fruit.

"Golden delicious" apples were dipped in 100 mg/L acibenzolar-S-methyl (ASM) to investigate the fruit quality and softening during 12 days of storage. Weight loss, flesh firmness, ethylene release, respiratory rate, content of total soluble solids and titratable acid, the activity of pectinase, cellulase, and ß-glucosidase, and water-insoluble pectin and water-soluble pectin contents were investigated. The results demonstrated that ASM treatment inhibited ethylene release and respiratory rate, reduced titratable acidity, and enhanced total soluble solids content in apples. Moreover, application of ASM suppressed the reduction of flesh firmness, activity of pectin methylesterase, and polygalacturonase. Cellulase, ß-glucosidase, and degradation of protopectin in apple fruit were also suppressed by ASM treatment during storage. In conclusion, ASM could maintain fruit quality by regulating cell wall-degrading enzymes during storage. PRACTICAL APPLICATIONS: Application of acibenzolar-S-methyl after harvest has the potential of delaying fruit softening by regulating cell wall-degrading enzymes, thus retain fruit quality.

The role of glucose-6-phosphate dehydrogenase in reactive oxygen species metabolism in apple exocarp induced by acibenzolar-S-methyl.

Apple exocarp was used to investigate the effect of acibenzolar-S-methyl (ASM) and dehydroepiandrosterone (DHEA) treatments on reaction oxygen species (ROS) metabolism. The results indicated that ASM enhanced the hydrogen peroxide (H2O2) content, the activities of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), glutathione reductase (GR) and glucose-6-phosphate dehydrogenase (G6PDH). ASM also increased the contents of ascorbic acid (AsA), reduced glutathione (GSH) and nicotinamide ademine dinucleotidephosphate (NADPH), MdSOD and MdAPX expression, but decreased MdMDHAR and dehydroascorbate reductase (MdDHAR) expression. DHEA suppressed H2O2 accumulation and POD, APX, MDHAR, G6PDH activities, but increased SOD, CAT and GR activities compared to the control. ASM and DHEA treatments suppressed the contents of AsA, GSH and NADPH, and expression of MdSOD, MdAPX and MdMDHAR. These results suggest that DHEA treatment prevented ROS metabolism induced by ASM which showed the important role of G6PDH in maintaining redox homeostasis in apple exocarp.

Theabrownin from Pu-erh tea attenuates hypercholesterolemia via modulation of gut microbiota and bile acid metabolism.

Pu-erh tea displays cholesterol-lowering properties, but the underlying mechanism has not been elucidated. Theabrownin is one of the most active and abundant pigments in Pu-erh tea. Here, we show that theabrownin alters the gut microbiota in mice and humans, predominantly suppressing microbes associated with bile-salt hydrolase (BSH) activity. Theabrownin increases the levels of ileal conjugated bile acids (BAs) which, in turn, inhibit the intestinal FXR-FGF15 signaling pathway, resulting in increased hepatic production and fecal excretion of BAs, reduced hepatic cholesterol, and decreased lipogenesis. The inhibition of intestinal FXR-FGF15 signaling is accompanied by increased gene expression of enzymes in the alternative BA synthetic pathway, production of hepatic chenodeoxycholic acid, activation of hepatic FXR, and hepatic lipolysis. Our results shed light into the mechanisms behind the cholesterol- and lipid-lowering effects of Pu-erh tea, and suggest that decreased intestinal BSH microbes and/or decreased FXR-FGF15 signaling may be potential anti-hypercholesterolemia and anti-hyperlipidemia therapies.

Effects of trisodium phosphate treatment after harvest on storage quality and sucrose metabolism in jujube fruit.

BACKGROUND: Trisodium phosphate (TSP), generally recognized as safe (GRAS), could control postharvest diseases and maintain fruit quality. However, changes of fruit quality and sucrose metabolism in harvested jujube after TSP treatment remain largely unknown. In the current study, jujube fruit (cv. sanxing) was used to study the effects of TSP on storage quality and sucrose metabolism during storage at 20 ± 2 °C with 40-50% relative humidity (RH). RESULTS: The results showed that 0.5 g L-1 TSP treatment reduced weight loss and reduced sugar content, suppressed the reduction of fruit firmness, maintained ascorbic acid (AsA) content and inhibited respiratory rate of jujube fruit. In addition, TSP treatment also reduced acid invertase (AI) and neutral invertase (NI) activities in sucrose metabolism in jujube fruit. Sucrose synthase-cleavage (SS-c), sucrose synthase-synthesis (SS-s) and sucrose phosphate synthase (SPS) activities were also suppressed by TSP treatment. CONCLUSION: Treatment with TSP could effectively reduce enzymes activities in sucrose metabolism and maintain storage quality of jujube fruit during storage. © 2019 Society of Chemical Industry.

Effect of sodium nitroprusside treatment on shikimate and phenylpropanoid pathways of apple fruit.

Blue mould caused by Penicillium expansum is one of the important diseases of apple fruit during storage. Phenylpropanoid pathway is an important induction mechanism that can utilize downstream metabolites of shikimate pathway to synthesize a series of secondary metabolites. Apple fruit (cv. Fuji) were treated with sodium nitroprusside (SNP) to study its effect on blue mould, shikimate and phenylpropanoid pathways. The results showed that 1.0 mmol L-1 SNP significantly inhibited lesion development of apple fruit inoculated with P. expansum. The results also indicated that SNP enhanced MdDHQS, MdSKDH, MdSK and MdEPSPS genes expressions, increased shikimic acid, tryptophan, tyrosine and phenylalanine contents in apple fruit. The activities of phenylalanine ammonialyase, 4-coumarate: coenzyme A, ligase, cinnamate 4-hydroxylase, lignin, total phenolic compounds and flavonoids contents in apple fruit were also increased by SNP treatment. These results suggest that SNP might modulate shikimate and phenylpropanoid pathways to enhance disease resistance of apple fruit.

Proton-Conductive Keggin-Type Clusters Decorated by the Complex Moieties of Cu(II) 2,2'-Bipyridine-4,4'-dicarboxylate/Diethyl Analogues.

Three proton-conductive decorated Keggin-type clusters, {[Cu(debpdc)(H2O)3][Cu(debpdc)(H2O)Cl][PMo12O40]} ·2CH3OH ·1.5CH3CN ·3H2O (1), {[Cu(H2bpdc)(H2O)2Cl0.5]2[PW12O40]}·10H2O (2), and {[Cu(H2bpdc)(H2O)2.5]2[SiW12O40]}·10H2O (3) (where debpdc is diethyl 2,2'-bipyridine-4,4'-dicarboxylate and H2bpdc is 2,2'-bipyridine-4,4'-dicarboxylic acid), were synthesized through electrostatic and coordination interactions between Keggin-type anions and Cu(II) H2bpdc/debpdc complex moieties. Interestingly, in the three complexes, both the H2bpdc/debpdc and the Keggin anion are covalently linked to the Cu2+ ions as polydentate organic and inorganic ligands, respectively. Notably, complexes 2 and 3 are the first examples of the functionalization of a Keggin-type cluster with Cu(II)-H2bpdc complex moieties, thereby providing a pathway to design and synthesize multifunctional hybrid materials with cluster structures based on two building units. In them, the free COOH groups of the H2bpdc ligand can act as both hydrogen bond acceptors and proton carriers. 1 has debpdc ligands with ethoxycarbonyl groups, while 2 and 3 have the H2bpdc ligands with free COOH groups; thus, the three complexes help us to understand the influence of the different substituents on the proton conductivity. The measurement results reveal that 2 and 3 have a high conductivity value of over 10-3 S cm-1 at 100 °C under 98% relative humidity, which is 2 orders of magnitude higher than that of 1 under the same conditions.

Application of sodium silicate retards apple softening by suppressing the activity of enzymes related to cell wall degradation.

BACKGROUND: During the storage of apples, apple softening is one of the main problems. Sodium silicate has been used to enhance disease resistance and maintain quality of fruits. In the present study, apple fruit (cv. Golden delicious) were treated with 100 mmol L-1 sodium silicate for 10 min and stored at 20 °C to investigate its effects on weight loss, flesh firmness, and the activity of cell wall-degrading enzymes. RESULTS: The results indicated that 100 mmol L-1 of sodium silicate treatment delayed the increase of weight loss and decrease of the flesh firmness in apples. Sodium silicate treatment also suppressed the activity of polygalacturonic acid transeliminase and pectin methyltranseliminase, pectin methylgalacturonase, polygalacturonase, cellulase and ß-galactosidase in the fruit. CONCLUSIONS: Delaying apple softening by sodium silicate treatment is closely related to the inhibition of the activity of cell wall-degrading enzymes and weight loss. © 2018 Society of Chemical Industry.

Changes in the sucrose metabolism in apple fruit following postharvest acibenzolar-S-methyl treatment.

BACKGROUND: Apple (cv. Ralls) fruit were treated with 0.1 g L-1 acibenzolar-S-methyl (ASM) for 10 min to evaluate the changes in enzyme activity and gene expression in the sucrose metabolism during storage at 20 °C with 30%-40% relative humidity. RESULTS: The results showed that sucrose phosphate synthase (SPS) and sucrose synthase synthesis (SS-s) activity was enhanced by ASM in apple fruit during the entire storage period. Sucrose synthase-cleavage (SS-c) and neutral invertase (NI) activity was suppressed by ASM treatment but acid invertase (AI) activity was increased in the middle period after ASM treatment. Acibenzolar-S-methyl treatment also significantly inhibited SPS and NI gene expression in apple fruit during storage. However, SS gene expression increased in the ASM-treated apple fruit. High levels of expression of the fructokinase (FK) and hexokinase (HK) genes were observed during the middle storage period in the ASM-treated fruit. CONCLUSION: Taken together, these results suggest that ASM delays the senescence of apple fruit by regulating the sugar metabolism. © 2018 Society of Chemical Industry.

Effect of trisodium phosphate dipping treatment on the quality and energy metabolism of apples.

The effects of postharvest trisodium phosphate (TSP) dipping (0.5 mg/mL) on the quality and mitochondrial energy metabolism of apple fruit (cv. Golden delicious) were studied. The results indicated that TSP treatment inhibited the respiration intensity, delayed the increase of weight loss, and inhibited the decrease of flesh firmness, ascorbic acid (AsA), titratable acid (TA) and soluble solids content (SSC) of apple fruit. The results also indicated that TSP treatment delayed the decline of the content of ATP, ADP and energy charge of apple fruit, and enhanced the activity of H+-ATPase, Ca2+-ATPase, succinate dehydrogenase and cytochrome C oxidase. These results suggested that TSP could maintain the quality of apple fruit by mediating respiration and mitochondrial energy metabolism.

Proton-Conductive Cocrystals of Twin Isomers of Coordination Polymers in Situ Formed by Keggin Anions and Cu(II)-4,4'-Bis(hydroxymethyl)-2,2'-bipyridine Complex Moieties.

Cocrystals and isomers, two well-known unique concepts in supramolecular chemistry, are rarely put together until now. For the first time, we report three unprecedented and interesting cocrystals of twin isomers of coordination polymers (CPs) in situ formed by typical Keggin anions and Cu(II)-4,4'-bis(hydroxymethyl)-2,2'-bipyridine (Cu(II)-H2L) complex moieties. In cocrystals 1-3, the Cu(II)-H2L complex moieties are quadrisupported on Keggin-type anions through W(Mo)-Ot-Cu-Ot-W(Mo) (Ot is the terminal O atom) links in the crystal to form two twin ionic/neutral CPs with a fixed chemical stoichiometry. Cocrystals 1-2 contain ionic isomers as {[Cu1(H2L)(H2O)2]2[P1W12O40]} n n+/{[Cu2(HL)(H2O)2]2[P2W12O40]} n n- for 1 and {[Cu1(H2L)(H2O)2]2[P1Mo12O40]} n n+/{[Cu2(HL)(H2O)2]2[P2Mo12O40]} n n- for 2. Cocrystal 3 contains neutral isomers as {[Cu1(H2L)(H2O)2]2[Si1W12O40]} n and {[Cu2(H2L)(H2O)2]2[Si2W12O40]} n. Cooperation of conformation and hydrogen bond network isomerism of Cu(II)-H2L fragments and tetracoordinated mode isomerism of Keggin anion is perfectly embodied in twin isomers. Moreover, based on hydrogen-bonding interactions, twin isomers are alternately arranged in a 1:1 stoichiometric ratio to give a cocrystal. Complicated accumulation of three types of hydrogen-bonding assemblies in the same crystal may be the reason that they give conductivity values over 10-4 S·cm-1 at 100 °C under 98% relative humidity.