The Effect of Maternal High-Fat or High-Carbohydrate Diet during Pregnancy and Lactation on Cytochrome P450 2D (CYP2D) in the Liver and Brain of Rat Offspring.

Cytochrome P450 2D (CYP2D) is important in psychopharmacology as it is engaged in the metabolism of drugs, neurosteroids and neurotransmitters. An unbalanced maternal diet during pregnancy and lactation can cause neurodevelopmental abnormalities and increases the offspring's predisposition to neuropsychiatric diseases. The aim of the present study was to evaluate the effect of maternal modified types of diet: a high-fat diet (HFD) and high-carbohydrate diet (HCD) during pregnancy and lactation on CYP2D in the liver and brain of male offspring at 28 (adolescent) or 63 postnatal days (young adult). The CYP2D activity and protein level were measured in the liver microsomes and the levels of mRNAs of CYP2D1, 2D2 and 2D4 were investigated both in the liver and brain. In the liver, both HFD and HCD increased the mRNA levels of all the three investigated CYP2D genes in adolescents, but an opposite effect was observed in young adults. The CYP2D protein level increased in adolescents but not in young adults. In contrast, young adults showed significantly decreased CYP2D activity. Similar effect of HFD on the CYP2D mRNAs was observed in the prefrontal cortex, while the effect of HCD was largely different than in the liver (the CYP2D2 expression was not affected, the CYP2D4 expression was decreased in young adults). In conclusion, modified maternal diets influence the expression of individual CYP2D1, CYP2D2 and CYP2D4 genes in the liver and brain of male offspring, which may affect the metabolism of CYP2D endogenous substrates and drugs and alter susceptibility to brain diseases and pharmacotherapy outcome.

A flexible linker of 8-amino acids between the membrane binding segment and the FMN domain of cytochrome P450 reductase is necessary for optimal activity.

The diflavin NADPH-cytochrome P450 reductase (CYPOR) plays a critical role in human cytochrome P450 (CYP) activity by sequentially delivering two electrons from NADPH to CYP enzymes during catalysis. Although electron transfer to forty-eight human CYP enzymes by the FMN hydroquinone of CYPOR is well-known, the role of the linker between the NH2-terminus membrane-binding domain (MBD) and FMN domain in supporting the activity of P450 enzymes remains poorly understood. Here we demonstrate that a linker with at least eight residues is required to form a functional CYPOR-CYP2B4 complex. The linker has been shortened in two amino-acid increments from Phe44 to Ile57 using site directed mutagenesis. The ability of the deletion mutants to support cytochrome P450 2B4 (CYP2B4) catalysis and reduce ferric CYP2B4 was determined using an in vitro assay and stopped-flow spectrophotometry. Steady-state enzyme kinetics showed that shortening the linker by 8-14 amino acids inhibited (63-99%) the ability of CYPOR to support CYP2B4 activity and significantly increased the Km of CYPOR for CYP2B4. In addition, the reductase mutants decreased the rate of reduction of ferric CYP2B4 (46-95%) compared to wildtype when the linker was shortened by 8-14 residues. These results indicate that a linker with a minimum length of eight residues is necessary to enable the FMN domain of reductase to interact with CYP2B4 to form a catalytically competent complex. Our study provides evidence that the length of the MBD-FMN domain linker is a major determinant of the ability of CYPOR to support CYP catalysis and drug metabolism by P450 enzymes. PREAMBLE: This manuscript is dedicated in memory of Dr. James R. Kincaid who was the doctoral advisor to Dr. Freeborn Rwere and a longtime collaborator and friend of Dr. Lucy Waskell. Dr. James R. Kincaid was a distinguished professor of chemistry specializing in resonance Raman (rR) studies of heme proteins. He inspired Dr. Rwere (a Zimbabwean native) and three other Zimbabweans (Dr. Remigio Usai, Dr. Daniel Kaluka and Ms. Munyaradzi E. Manyumwa) to use lasers to document subtle changes occurring at heme active site of globin proteins (myoglobin and hemoglobin) and cytochrome P450 enzymes. Dr. Rwere appreciate his contributions to the development of talented Black scientists from Africa.

Pregnane X receptor activation induces liver enlargement and regeneration and simultaneously promotes the metabolic activity of CYP3A1/2 and CYP2C6/11 in rats.

Human pregnane X receptor (PXR) is critical for regulating the expression of key drug-metabolizing enzymes such as CYP3A and CYP2C. Our recent study revealed that treatment with rodent-specific PXR agonist pregnenolone-16α-carbonitrile (PCN) significantly induced hepatomegaly and promoted liver regeneration after two-thirds partial hepatectomy (PHx) in mice. However, it remains unclear whether PXR activation induces hepatomegaly and liver regeneration and simultaneously promotes metabolic function of the liver. Here, we investigated the metabolism activity of CYP1A2, CYP3A1/2 and CYP2C6/11 during PXR activation-induced liver enlargement and regeneration in rats after cocktail dosing of CYP probe drugs. For PCN-induced hepatomegaly, a notable increase in the metabolic activity of CYP3A1/2 and CYP2C6/11, as evidenced by the plasma exposure of probe substrates and the AUC ratios of the characteristic metabolites to its corresponding probe substrates. The metabolic activity of CYP1A2, CYP3A1/2 and CYP2C6/11 decreased significantly after PHx. However, PCN treatment obviously enhanced the metabolic activity of CYP2C6/11 and CYP3A1/2 in PHx rats. Furthermore, the protein expression levels of CYP3A1/2 and CYP2C6/11 in liver were up-regulated. Taken together, this study demonstrates that PXR activation not only induces hepatomegaly and liver regeneration in rats, but also promotes the protein expression and metabolic activity of the PXR downstream metabolizing enzymes such as CYP3A1/2 and CYP2C6/11 in the body.

Relationship between Vitamin D3 Deficiency, Metabolic Syndrome and VDR, GC, and CYP2R1 Gene Polymorphisms.

The presence of vitamin D3 deficiency associated with the presence of metabolic syndrome (MS) has important public health effects. This study aims to investigate the relationship between vitamin D3 deficiency, MS and vitamin D3 receptor (VDR), GC Vitamin D binding protein (GC), and cytochrome P450 family 2 subfamily R member 1 (CYP2R1) gene polymorphisms, and genes whose encoded proteins are responsible for vitamin D3 metabolism and transport. A total of 58 participants were included in this study (age 39 ± 12 years) and were selected over a 12-month period. They were divided into four groups, depending on the presence of polymorphisms in VDR, GC, and CYP2R1 genes and their weight status. At baseline, in months 3, 6, and 12, biochemical parameters including 25(OH)D3, total cholesterol, LDL cholesterol, HDL cholesterol, triglycerides, and homeostatic model assessment (HOMA index), the insulin resistance indicator were measured. Our results show that all subjects in the polymorphism group supplemented with vitamin D3 reached an optimal level of vitamin D3 associated with high concentrations of 25(OH)D3. Weight loss was most significant in patients in the POW group (overweight patients).

Epoxygenase Cyp2c44 Regulates Hepatic Lipid Metabolism and Insulin Signaling by Controlling FATP2 Localization and Activation of the DAG/PKCδ Axis.

Cytochrome P450 epoxygenase Cyp2c44, a murine epoxyeicosatrienoic acid (EET)-producing enzyme, promotes insulin sensitivity, and Cyp2c44-/- mice show hepatic insulin resistance. Because insulin resistance leads to hepatic lipid accumulation and hyperlipidemia, we hypothesized that Cyp2c44 regulates hepatic lipid metabolism. Standard chow diet (SCD)-fed male Cyp2c44-/- mice had significantly decreased EET levels and increased hepatic and plasma lipid levels compared with wild-type mice. We showed increased hepatic plasma membrane localization of the FA transporter 2 (FATP2) and total unsaturated fatty acids and diacylglycerol (DAG) levels. Cyp2c44-/- mice had impaired glucose tolerance and increased hepatic plasma membrane-associated PKCδ and phosphorylated IRS-1, two negative regulators of insulin signaling. Surprisingly, SCD and high-fat diet (HFD)-fed Cyp2c44-/- mice had similar glucose tolerance and hepatic plasma membrane PKCδ levels, suggesting that SCD-fed Cyp2c44-/- mice have reached their maximal glucose intolerance. Inhibition of PKCδ resulted in decreased IRS-1 serine phosphorylation and improved insulin-mediated signaling in Cyp2c44-/- hepatocytes. Finally, Cyp2c44-/- HFD-fed mice treated with the analog EET-A showed decreased hepatic plasma membrane FATP2 and PCKδ levels with improved glucose tolerance and insulin signaling. In conclusion, loss of Cyp2c44 with concomitant decreased EET levels leads to increased hepatic FATP2 plasma membrane localization, DAG accumulation, and PKCδ-mediated attenuation of insulin signaling. Thus, Cyp2c44 acts as a regulator of lipid metabolism by linking it to insulin signaling.

Suppression of hepatic ChREBP⍺-CYP2C50 axis-driven fatty acid oxidation sensitizes mice to diet-induced MASLD/MASH.

OBJECTIVES: Compromised hepatic fatty acid oxidation (FAO) has been observed in human MASH patients and animal models of MASLD/MASH. It remains poorly understood how and when the hepatic FAO pathway is suppressed during the progression of MASLD towards MASH. Hepatic ChREBP⍺ is a classical lipogenic transcription factor that responds to the intake of dietary sugars. METHODS: We examined its role in regulating hepatocyte fatty acid oxidation (FAO) and the impact of hepatic Chrebpa deficiency on sensitivity to diet-induced MASLD/MASH in mice. RESULTS: We discovered that hepatocyte ChREBP⍺ is both necessary and sufficient to maintain FAO in a cell-autonomous manner independently of its DNA-binding activity. Supplementation of synthetic PPAR⍺/δ agonist is sufficient to restore FAO in Chrebp-/- primary mouse hepatocytes. Hepatic ChREBP⍺ was decreased in mouse models of diet-induced MAFSLD/MASH and in patients with MASH. Hepatocyte-specific Chrebp⍺ knockout impaired FAO, aggravated liver steatosis and inflammation, leading to early-onset fibrosis in response to diet-induced MASH. Conversely, liver overexpression of ChREBP⍺-WT or its non-lipogenic mutant enhanced FAO, reduced lipid deposition, and alleviated liver injury, inflammation, and fibrosis. RNA-seq analysis identified the CYP450 epoxygenase (CYP2C50) pathway of arachidonic acid metabolism as a novel target of ChREBP⍺. Over-expression of CYP2C50 partially restores hepatic FAO in primary hepatocytes with Chrebp⍺ deficiency and attenuates preexisting MASH in the livers of hepatocyte-specific Chrebp⍺-deleted mice. CONCLUSIONS: Our findings support the protective role of hepatocyte ChREBPa against diet-induced MASLD/MASH in mouse models in part via promoting CYP2C50-driven FAO.

Gene expression profiling of vitamin D metabolism enzymes in leukemia and lymphoma patients: molecular aspect interplay of VDR, CYP2R1, and CYP24A1.

BACKGROUND: Vitamin D deficiency is prevalent among the Indonesian population, particularly in individuals diagnosed with leukemia-lymphoma. The regulation of vitamin D metabolism is influenced by the expression of several enzymes, such as CYP2R1, CYP24A1, and the vitamin D receptor (VDR). This study aimed to scrutinize the gene expression profiles in both mRNA and protein levels of VDR, CYP2R1, and CYP24A1 in leukemia and lymphoma patients. METHOD: The research was a cross-sectional study conducted at Cipto Mangunkusumo Hospital (RSCM) in Jakarta, Indonesia. The study included a total of 45 patients aged over 18 years old who have received a diagnosis of lymphoma or leukemia. Vitamin D status was measured by examining serum 25 (OH) D levels. The analysis of VDR, CYP2R1, and CYP24A1 mRNA expression utilized the qRT-PCR method, while protein levels were measured through the ELISA method. CONCLUSION: The study revealed a noteworthy difference in VDR protein levels between men and women. The highest mean CYP24A1 protein levels were observed in the age group > 60 years. This study found a significant, moderately positive correlation between VDR protein levels and CYP24A1 protein levels in the male and vitamin D sufficiency groups. In addition, a significant positive correlation was found between VDR mRNA levels and CYP2R1 mRNA levels, VDR mRNA levels and CYP2R1 mRNA levels, and CYP2R1 mRNA levels and CYP24A1 mRNA levels. However, the expression of these genes does not correlate with the protein levels of its mRNA translation products in blood circulation.

Decreased vitamin D bio-availability with altered DNA methylation of its metabolism genes in association with the metabolic disorders among the school-aged children with degree I, II, and III obesity.

Obesity is strongly associated with disturbances of vitamin D (VD) metabolites in the animal models. However, the related epidemiological evidence is still controversial, especially the different degrees of obesity children. Hence, in this present representative case-control study, 106 obesity school-age children aged 7-12 years were included and divided into different subgroups as degree I (the age- and sex-specific BMI≥95th percentile, n=45), II (BMI ≥120% percentile, n=34) and III (BMI ≥140% percentile, n=27) obesity groups across the ranges of body mass index (BMI). While the age- and sex-matched subjects without obesity were as the control group. Notably, it was significantly different of body composition, anthropological and clinical characteristics among the above four subgroups with the dose-response relationships (P<.05). Moreover, comparing with the control group, the serum VD concentrations were higher, VD metabolites like 25(OH)D, 25(OH)D3 and 1,25(OH)2D, and related hydroxylases as CYP27A1, CYP2R1 and CYP27B1 were lower in the degree I, II, and III obesity subgroups (P<.05), which were more disorder with the anthropological and clinical characteristics as the obesity was worsen in a BMI-independent manner (P<.05). However, there was a significant increase of CYP27B1 in the degree III obesity group than those in the degree I and II obesity subgroups. Furthermore, the methylation patterns on the genome-wide (Methylation/Hydroxymethylation) and VD metabolism genes (CYP27A1, CYP2R1 and CYP27B1) were negatively correlated with the worse obesity and their related expressions (P<.05). In summary, these results indicated that obesity could affect the homeostasis of VD metabolism related genes such as CYP27A1, CYP2R1, CYP27B1 and etc through abnormal DNA methylation, resulting in the disorders of VD related metabolites to decrease VD bio-availability with the BMI-independent manner. In turn, the lower levels of VD metabolites would affect the liver function to exacerbate the progression of obesity, as the Degree II and III obesity subgroups.

CircPHKB decreases the sensitivity of liver cancer cells to sorafenib via miR-1234-3p/CYP2W1 axis.

Sorafenib is currently the first-line treatment for patients with advanced liver cancer, but its therapeutic efficacy declines significantly after a few months of treatment. Therefore, it is of great importance to investigate the regulatory mechanisms of sorafenib sensitivity in liver cancer cells. In this study, we provided initial evidence demonstrating that circPHKB, a novel circRNA markedly overexpressed in sorafenib-treated liver cancer cells, attenuated the sensitivity of liver cancer cells to sorafenib. Mechanically, circPHKB sequestered miR-1234-3p, resulting in the up-regulation of cytochrome P450 family 2 subfamily W member 1 (CYP2W1), thereby reducing the killing effect of sorafenib on liver cancer cells. Moreover, knockdown of circPHKB sensitized liver cancer cells to sorafenib in vivo. The findings reveal a novel circPHKB/miR-1234-3p/CYP2W1 pathway that decreases the sensitivity of liver cancer cells to sorafenib, suggesting that circPHKB and the axis may serve as promising targets to improve the therapeutic efficacy of sorafenib against liver cancer.

Single nucleotide polymorphisms in vitamin D binding protein and 25-hydroxylase genes affect vitamin D levels in adolescents of Arab ethnicity in Kuwait.

Vitamin D deficiency (VDD) is widespread in the Arab world despite ample sunshine throughout the year. In our previous study, lifestyle and socio-demographic factors could explain only 45% of variability in vitamin D levels in Kuwaiti adolescents, suggesting that genetics might contribute to VDD in this region. Single nucleotide polymorphisms (SNP) in the 25-hydroxylase (CYP2R1) and the GC globulin (GC) genes have been reported to affect vitamin D levels in various ethnic groups in adults. In this study, we investigated the association of two SNPs from GC (rs4588 and rs7041) and three SNPs from CYP2R1 (rs10741657, rs11023374 and rs12794714) with vitamin D levels and VDD in a nationally representative sample of adolescents of Arab ethnicity from Kuwait. Multivariable linear regression, corrected for age, sex, parental education, governorate, body mass index, and exposure to sun, demonstrated that each of the 5 study variants showed significant associations with plasma 25(OH)D levels in one or more of the additive, recessive, and dominant genetic models - the rs10741657 under all the three models, rs12794714 under both the additive and recessive models, rs7041 under the recessive model; and rs4588 and rs11023374 under the dominant model. Minor alleles at rs4588 (T), rs7041 (A), rs11023374 (C), and rs12794714 (A) led to a decrease in plasma 25(OH)D levels - rs4588:[ß (95%CI) = -4.522 (-8.66,-0.38); p=0.033]; rs7041:[ß (95%CI) = -6.139 (-11.12,-1.15); p=0.016]; rs11023374:[ß (95%CI) = -4.296 (-8.18,-0.40); p=0.031]; and rs12794714:[ß (95%CI) = -3.498 (-6.27,-0.72); p=0.014]. Minor allele A at rs10741657 was associated with higher levels of plasma 25(OH)D levels [ß (95%CI) = 4.844 (1.62,8.06); p=0.003)] and lower odds of vitamin D deficiency (OR 0.40; p=0.002). These results suggest that the CYP2R1 and GC SNP variants are partly responsible for the high prevalence of VDD in Kuwait. Genotyping these variants may be considered for the prognosis of VDD in Kuwait.

CRY1/2 regulate rhythmic CYP2A5 in mouse liver through repression of E4BP4.

CYP2A5, an enzyme responsible for metabolism of diverse drugs, displays circadian rhythms in its expression and activity. However, the underlying mechanisms are not fully established. Here we aimed to investigate a potential role of CRY1/2 (circadian clock modulators) in circadian regulation of hepatic CYP2A5. Regulatory effects of CRY1/2 on CYP2A5 were determined using Cry1-null and Cry2-null mice, and validated using AML-12, Hepa1-6 and HepG2 cells. CYP2A5 activities both in vivo and in vitro were assessed using coumarin 7-hydroxylation as a probe reaction. mRNA and protein levels were detected by qPCR and western blotting, respectively. Regulatory mechanism was studied using a combination of luciferase reporter assays, chromatin immunoprecipitation (ChIP) and co-immunoprecipitation (Co-IP). We found that ablation of Cry1 or Cry2 in mice reduced hepatic CYP2A5 expression (at both mRNA and protein levels) and blunted its diurnal rhythms. Consistently, these knockouts showed decreased CYP2A5 activity (characterised by coumarin 7-hydroxylation) and a loss of its time-dependency, as well as exacerbated coumarin-induced hepatotoxicity. Cell-based assays confirmed that CRY1/2 positively regulated CYP2A5 expression and rhythms. Based on combined luciferase reporter, ChIP and Co-IP assays, we unraveled that CRY1/2 interacted with E4BP4 protein to repress its inhibitory effect on Cyp2a5 transcription and expression. In conclusion, CRY1/2 regulate rhythmic CYP2A5 in mouse liver through repression of E4BP4. These findings advance our understanding of circadian regulation of drug metabolism and pharmacokinetics.

Association of Single Nucleotide Polymorphism in VDR, GC Globulin and CYP2R1 with the Risk of Esophageal Cancer.

BACKGROUND: The proactive role of vitamin D has been well determined in different cancers. The protein that encodes the components of the vitamin D metabolism could appear to play a pivotal role in vitamin D stability and its maintenance. A polymorphism in vitamin-D-receptor (VDR), carrier globulin/binding protein (GC) and cytochrome P-450 family 2, subfamily R, polypeptide 1 (CYP2R1) genes has been predicted to be associated with the development of cancer. This study was designed to detect the association of VDR, GC Globulin and CYP2R1 gene polymorphism with the risk of esophageal cancer in the North-east Indian population. METHODS: To carry out the study, a total of 100 patients diagnosed with esophageal cancer and 101 healthy controls were enrolled. In a case-control manner, all samples were subjected to do genotype testing for known SNPs on the VDR (rs1544410), GC (rs4588), and CYP2R1 (rs10741657) genes using Restriction-fragment length polymorphism (RFLP) followed by Sanger sequencing. The collected demographic and clinical data were analysed using the statistical software package SPSS v22.0. RESULTS: The VDR haplotype heterozygous TC was found strongly associated with the carcinoma group (OR:1.09, 95%CI:0.67-1.75). The risk factors analysis using the GC globulin rs4588 phenotype, found a positive correlation in terms of mutant AA's harmful influence on the cancer cohort (OR = 1.125, OR=1.125, 95% CI, 0.573-2.206). The influence of the CYP2R1 rs10741657 polymorphism on the malignant cohort revealed that the GG mutant had a significant negative influence on the carcinoma, has an influential role in disease severity ( OR:1.736, at 95% CI; 0.368-8.180). CONCLUSION: In conclusion, this study revealed the potential association of VDR gene polymorphism in the progression and development of esophageal cancer in north east Indian population cohort.

Correlation of CYP2R1 gene promoter methylation with circulating vitamin D levels among healthy adults.

Background & objectives: Despite being a tropical country, vitamin D deficiency is highly prevalent in India with studies indicating 40-99 per cent prevalence. Apart from calcium and phosphate metabolism, vitamin D is involved in cell cycle regulation, cardiovascular, hepatoprotection. The metabolism of vitamin D is regulated by vitamin D tool genes (CYP2R1/CYP27B1/CYP24A1/VDR). The promoter regions of some of these genes have CpG islands, making them prone to methylation induced gene silencing, which may cause a reduction in circulating vitamin D levels. Epigenetic basis of vitamin D deficiency is yet to be studied in India, and hence, this pilot study was aimed to analyze whether methylation levels of CYP2R1 gene were correlated with the levels of 25(OH)D in healthy, adult individuals in Indian population. Methods: In this cross-sectional study, healthy adults of 18-45 yr of age with no history of malabsorption, thyroidectomy, chronic illness or therapeutic vitamin D supplementation were recruited. DNA methylation analysis was carried out by methylation specific quantitative PCR. Serum calcium, phosphate and vitamin D levels were also quantified. Statistical analysis was done by R 4.0.5 software. Results: A total of 61 apparently healthy adults were analyzed. The serum vitamin D levels did not correlate with CYP2R1 methylation levels in our study population. Significant positive correlation was observed between age and serum vitamin D levels. Significant association of gender was found with CYP2R1 methylation levels. Interpretation & conclusions: This study found no significant correlation between levels of CYP2R1 methylation and circulating 25(OH)D deficiency. Further studies on the Indian population having a larger sample size including entire vitamin D tool genes, among different ethnic groups may be conducted to elucidate molecular etiology of circulating 25(OH)D deficiency. The high prevalence of normal serum calcium and phosphate levels among vitamin D deficient subjects in this study coupled with the strikingly high prevalence of the deficiency at the national level, may suggest the need to revise the cut-off criteria for vitamin D deficiency in the Indian population.

Genetic variants in the vitamin D pathway and their association with vitamin D metabolite levels: Detailed studies of an inner-city pediatric population suggest a modest but significant effect in early childhood.

OBJECTIVES: In a large cohort of healthy infants and toddlers 6-36 months of age (n = 776), we have been exploring the potential role of genetic variation in predisposition to vitamin D insufficiency. The genes encoding the key cytochrome P450 hydroxylases (CYP2R1, CYP24A1, and CYP27B1) harbour recurrent mutations of uncertain effect. This study was undertaken to look for biochemically relevant associations of these variants with inter-individual differences in vitamin D metabolism in an at-risk pediatric population. METHODS: Genotyping for CYP2R1-CT (c.-1127 C>T, rs10741657), CYP24A1-AG (c.-686A>G, rs111622401), and CYP27B1-CA (c.-1261 C>A, rs10877012) mutations were performed using SNaPshot assay, followed by Sanger sequencing confirmation. Vitamin D metabolites and vitamin D binding protein (DBP) were measured by established methods. RESULTS: In a multivariate regression model, with corrections for co-variates, subjects with the homozygous CYP2R1-TT variant had significantly higher concentrations of 25(OH)D, free 25(OH)D, and 24,25(OH)2D levels. In subjects with the CYP24A1-AG mutation, concentrations of 25(OH)D were significantly higher. CONCLUSIONS: The CYP2R1-TT and CYP24A1-AG variants have measurable effects on the vitamin D pathway. It seems unlikely that they will be clinically relevant in isolation, but they may be members of the large pool of infrequent mutations contributing to different risks for the vitamin D deficiency phenotype.

Leptin and IGF-1 in Infancy Are Associated With Variants in DHCR7 and CYP2R1 That Relate With Type 1 Diabetes and 25OHD.

CONTEXT: Vitamin D has been variably implicated in risk of developing type 1 diabetes based on cohorts of at-risk individuals. Emergent type 1 diabetes in childhood is putatively preceded by altered growth. OBJECTIVE: We explored whether polymorphisms in vitamin D metabolism genes modify risk of type 1 diabetes via effects on growth in a prospective, population-based cohort of infants. METHODS: The Cambridge Baby Growth Study enrolled newborns from Cambridgeshire, UK, for follow-up in infancy. In 612 infants, we genotyped single nucleotide polymorphisms in vitamin D metabolism genes that relate with type 1 diabetes: rs10741657 and rs12794714 in CYP2R1, rs12785878 in DHCR7, and rs10877012 in CYP27B1. Multivariate linear regression analyses tested associations between genotypes and anthropometric indices (weight, length, and skinfold thickness) or growth-related hormones (C-peptide, IGF-1, and leptin) in infancy. RESULTS: Birth weight showed borderline associations with the diabetes risk-increasing alleles in CYP2R1, rs10741657 (ß = -.11, P = .02) and rs12794714 (ß = -.09, P = .04). The risk-increasing allele rs12794714 was also associated with higher IGF-1 levels at age 24 months (ß = .30, P = .01). At age 3 months, the risk-increasing allele rs12785878 in DHCR7, known to negatively associate with 25-hydroxyvitamin D levels, showed a positive association with leptin levels (ß = .23, P = .009), which was pronounced in girls (P = .004) vs boys (P = .7). CONCLUSION: The vitamin D metabolism genes DHCR7 and CYP2R1 might influence infancy leptin and IGF-1 levels respectively. These findings open the possibility for a developmental role of vitamin D that is mediated by growth-related hormones with implications for the onset of type 1 diabetes autoimmunity.

[Analysis of CYP2U1 gene variants in a child with Hereditary spastic paraplegia type 56].

OBJECTIVE: To analyze the clinical phenotype and genetic characteristics of a child with Hereditary spastic paraplegia (HSP). METHODS: A child with HSP who was admitted to the Third Affiliated Hospital of Zhengzhou University on August 10, 2020 due to discovery of tiptoeing for 2 years was selected as the study subject, and relevant clinical data was collected. Peripheral blood samples of the child and her parents were collected for the extraction of genomic DNA. And trio-whole exome sequencing (trio-WES) was carried out. Candidate variants were verified by Sanger sequencing. Bioinformatic software was used to analyze the conservation of variant sites. RESULTS: The child was a 2-year-and-10-month-old female with clinical manifestations including increased muscle tone of lower limbs, pointed feet, and cognitive language delay. Trio-WES results showed that she had harbored compound heterozygous variants of c.865C>T (p.Gln289*) and c.1126G>A (p.Glu376Lys) of the CYP2U1 gene. And the corresponding amino acid for c.1126G>A (p.Glu376Lys) is highly conserved among various species. Based on guidelines from the American College of Medical Genetics and Genomics, the c.865C>T was predicted as a pathogenic variant (PVS1+PM2_Supporting), and c.1126G>A was rated as a variant of uncertain significance (PM2_Supporting+PM3+PP3). CONCLUSION: The child was diagnosed with HSP type 56 due to compound variants of the CYP2U1 gene. Above findings have enriched the mutation spectrum of the CYP2U1 gene.

Downregulation of the gene expression of Cyp2c29 and Cyp3a11 by cecal ligation and puncture-induced sepsis is associated with interleukin-6.

Sepsis is a pathological condition that affects the metabolism of administered drugs, leading to changes in the duration and intensity of their intended efficacies. Proinflammatory cytokines downregulate the expression of cytochrome P450s (P450s). The effects of P450 expression under inflammatory conditions have been studied using prophlogistic substances such as lipopolysaccharide; however, few studies have focused on clinical models of sepsis. Here, we show that cecal ligation and puncture (CLP), an approach for the study of human polymicrobial sepsis, leads to the expression of interleukin-1ß (IL-1ß), IL-6, and tumor necrosis factor α (TNFα) at 24 h after the CLP operation. Following CLP, IL-6-/- mice exhibited markedly lower survival than WT mice. In addition, CLP led to the significant downregulation of Cyp2c29 and Cyp3a11 gene expression in IL-1α-/-/ß-/- (IL-1-/-) and TNFα-/- mice as well as in WT mice. In contrast, CLP elicited no significant effect on Cyp3a11 expression in IL-6-/- mice. Although CLP reduced the Cyp2c29 expression level in IL-6-/- mice, the expression of Cyp2c29 was lower in CLP-operated WT mice than in CLP-operated IL-6-/- mice. The reduction in the respective P450 protein levels and activities due to CLP-induced sepsis, reflected in the mRNA expression levels, was abolished by IL-6 depletion. Thus, CLP-induced sepsis downregulates P450 gene expression, particularly Cyp2c expression, and this effect is associated with IL-6 without affecting resistance to CLP-induced sepsis. These findings demonstrate the usefulness of CLP for studying the regulation of P450s and highlight IL-6 as a potential indicator of drug-metabolizing capacity under septic conditions.

Polymorphisms in VDR, CYP27B1, CYP2R1, GC and CYP24A1 Genes as Biomarkers of Survival in Non-Small Cell Lung Cancer: A Systematic Review.

The objective of this systematic review was to provide a compilation of all the literature available on the association between single-nucleotide polymorphisms (SNPs) in the genes involved in the metabolic pathway of vitamin D and overall survival (OS) and progression-free survival (PFS) in patients with non-small cell lung cancer (NSCLC). This systematic review was conducted in accordance with the PRISMA guidelines. It included all the literature published up to 1 November 2022 and was carried out in four databases (Medline [PubMed], Scopus, Web of Science, and Embase), using the PICO strategy, with relevant keywords related to the objective. The quality of the studies included was evaluated with an assessment tool derived from the Strengthening the Reporting of Genetic Association Studies (STREGA) statement. Six studies were included in this systematic review. Our findings showed that the BsmI (rs1544410), Cdx-2 (rs11568820), FokI (rs2228570), ApaI (rs7975232), TaqI (rs731236), rs4646536, rs6068816, rs7041, and rs10741657 SNPs in the genes that play a part in vitamin D synthesis (CYP2R1, CYP27B1), transport (GC), and metabolism (CYP24A1), as well as in the vitamin D receptor (VDR), are associated with OS and/or PFS in patients with NSCLC. The SNPs in VDR have been the most extensively analyzed. This systematic review summed up the available evidence concerning the association between 13 SNPs in the main genes involved in the vitamin D metabolic pathway and prognosis in NSCLC. It revealed that SNPs in the VDR, CYP27B1, CYP24A1, GC, and CYP2R1 genes could have an impact on survival in this disease. These findings suggest the identification of prognostic biomarkers in NSCLC patients. However, evidence remains sparse for each of the polymorphisms examined, so these findings should be treated with caution.

Activated Hepatic Nuclear Factor-κB in Experimental Colitis Regulates CYP2A5 and Metronidazole Disposition.

Systemic exposure of metronidazole is increased in patients with inflammatory bowel diseases (IBDs), while the underlying mechanism remains unknown. Here, we aim to decipher the mechanisms by which experimental colitis regulates metronidazole disposition in mice. We first confirmed that the systemic exposure of metronidazole was elevated in dextran sulfate sodium (DSS)-induced experimental colitis. Hepatic microsomal incubation with metronidazole revealed that the production rate of 2-hydroxymetronidazole was inhibited, suggestive of a diminished hydroxylation reaction upon colitis. Remarkably, the hydroxylation reaction of metronidazole was selectively catalyzed by CYP2A5, which was downregulated in the liver of colitis mice. In addition, hepatic nuclear factor (NF)-κB (a prototypical and critical signaling pathway in inflammation) was activated in colitis mice. Luciferase reporter and chromatin immunoprecipitation assay indicated that NF-κB downregulated Cyp2a5 transcription through binding to an NF-κB binding site (-1711 to -1720 bp) in the promoter. We further verified that the regulatory effects of colitis on CYP2A5 depended on the disease itself rather than the DSS compound. First, one-day administration of DSS did not alter mRNA and protein levels of CYP2A5. Moreover, CYP2A5 was suppressed in the Il-10-/- spontaneously developing colitis model. Furthermore, Cyp2a5 expression was downregulated in both groups of mice with modest or severe colitis, whereas the expression change was much more significant in severe colitis as compared to modest colitis. Altogether, activated hepatic NF-κB in experimental colitis regulates CYP2A5 and metronidazole disposition, revealing the mechanism of pharmacokinetic instability under IBDs, and providing a theoretical foundation for rational drug use in the future.

Analysis of CYP2U1 gene variants in a child with Hereditary spastic paraplegia type 56 / 中华医学遗传学杂志

OBJECTIVE@#To analyze the clinical phenotype and genetic characteristics of a child with Hereditary spastic paraplegia (HSP).@*METHODS@#A child with HSP who was admitted to the Third Affiliated Hospital of Zhengzhou University on August 10, 2020 due to discovery of tiptoeing for 2 years was selected as the study subject, and relevant clinical data was collected. Peripheral blood samples of the child and her parents were collected for the extraction of genomic DNA. And trio-whole exome sequencing (trio-WES) was carried out. Candidate variants were verified by Sanger sequencing. Bioinformatic software was used to analyze the conservation of variant sites.@*RESULTS@#The child was a 2-year-and-10-month-old female with clinical manifestations including increased muscle tone of lower limbs, pointed feet, and cognitive language delay. Trio-WES results showed that she had harbored compound heterozygous variants of c.865C>T (p.Gln289*) and c.1126G>A (p.Glu376Lys) of the CYP2U1 gene. And the corresponding amino acid for c.1126G>A (p.Glu376Lys) is highly conserved among various species. Based on guidelines from the American College of Medical Genetics and Genomics, the c.865C>T was predicted as a pathogenic variant (PVS1+PM2_Supporting), and c.1126G>A was rated as a variant of uncertain significance (PM2_Supporting+PM3+PP3).@*CONCLUSION@#The child was diagnosed with HSP type 56 due to compound variants of the CYP2U1 gene. Above findings have enriched the mutation spectrum of the CYP2U1 gene.