Postoperative jaundice related to UGT1A1 and ABCB11 gene mutations: A case report and literature review.

BACKGROUND: Patients with obstructive jaundice caused by intrahepatic bile duct stones can be effectively managed by surgery. However, some patients may develop postoperative complications, liver failure, and other life-threatening situations. Here, we report a patient with mutations in the uridine 5'-diphospho-glucuronosyltransferase 1A1 (UGT1A1) and bile salt export pump (adenosine triphosphate-binding cassette subfamily B member 11, ABCB11) genes who presented multiple intrahepatic bile duct stones and cholestasis, and the jaundice of the patient increased after partial hepatectomy. CASE SUMMARY: A 52-year-old male patient admitted to the hospital on October 23, 2021, with a progressive exacerbation of jaundice, was found to have multiple intrahepatic bile duct stones with the diagnoses of obstructive jaundice and acute cholecystitis. Subsequently, the patient underwent left hepatectomy with biliary exploration, stone extraction, T-tube drainage, and cholecystectomy without developing any intraoperative complications. The patient had a dark urine color with worsening jaundice postoperatively and did not respond well to plasma exchange and other symptomatic and supportive treatments. Since the progressive increase in postoperative bilirubin could not be clinically explained with any potential reason, including, if not at all, viral infection, cholangitis, autoimmune liver disease, and other causes, the patient underwent whole-exon screening for any genetic diseases, which surprisingly identified UGT1A1 and ABCB11 gene mutations related to glucuronidation of indirect bilirubin as well as bile acid transport in hepatocytes, respectively. Thus, we hypothesized that postoperative refractory cholestasis might result from UGT1A1 and ABCB11 gene mutations and further recommended liver transplantation to the patient, who eventually declined it and died from liver failure six months later. CONCLUSION: Surgery may aggravate cholestasis in patients with multiple intrahepatic bile duct stones and cholestasis associated with UGT1A1 and ABCB11 gene mutations. A liver transplant may be the best option if active medical treatment fails.

Feedback loop between hepatocyte nuclear factor 1α and endoplasmic reticulum stress mitigates liver injury by downregulating hepatocyte apoptosis.

Hepatocyte nuclear factor alpha (HNF1α), endoplasmic reticulum (ER) stress, and hepatocyte apoptosis contribute to severe acute exacerbation (SAE) of liver injury. Here, we explore HNF1α-ER stress-hepatocyte apoptosis interaction in liver injury. LO2, HepG2 and SK-Hep1 cells were treated with thapsigargin (TG) or tunicamycin (TM) to induce ER stress. Carbon tetrachloride (CCl4) was used to induce acute liver injury in mice. Low-dose lipopolysaccharide (LPS) exacerbated liver injury in CCl4-induced mice. Significant apoptosis, HNF1α upregulation, and nuclear factor kappa B (NF-κB) activation were observed in human-derived hepatocytes during ER stress. Knockdown of Rela, NF-κB p65, inhibited the HNF1α upregulation. Following CCl4 treatment ER stress, apoptosis, HNF1α expression and RelA phosphorylation were significantly increased in mice. HNF1α knockdown reduced activating transcription factor 4 (ATF4) expression, and aggravated ER stress as well as hepatocyte apoptosis in vivo and in vitro. The double fluorescent reporter gene assay confirmed that HNF1α regulated the transcription of ATF4 promoter. LPS aggravated CCl4-induced liver injury and reduced HNF1α, and ATF4 expression. Therefore, in combination, HNF1α and ER stress could be mutually regulated forming a feedback loop, which helps in protecting the injured liver by down-regulating hepatocyte apoptosis. Low-dose LPS aggravates hepatocyte apoptosis and promotes the SAE of liver injury by interfering with the feedback regulation of HNF1α and ER stress in acute liver injury.

Mitochondrial FAD shortage in SLC25A32 deficiency affects folate-mediated one-carbon metabolism.

The SLC25A32 dysfunction is associated with neural tube defects (NTDs) and exercise intolerance, but very little is known about disease-specific mechanisms due to a paucity of animal models. Here, we generated homozygous (Slc25a32Y174C/Y174C and Slc25a32K235R/K235R) and compound heterozygous (Slc25a32Y174C/K235R) knock-in mice by mimicking the missense mutations identified from our patient. A homozygous knock-out (Slc25a32-/-) mouse was also generated. The Slc25a32K235R/K235R and Slc25a32Y174C/K235R mice presented with mild motor impairment and recapitulated the biochemical disturbances of the patient. While Slc25a32-/- mice die in utero with NTDs. None of the Slc25a32 mutations hindered the mitochondrial uptake of folate. Instead, the mitochondrial uptake of flavin adenine dinucleotide (FAD) was specifically blocked by Slc25a32Y174C/K235R, Slc25a32K235R/K235R, and Slc25a32-/- mutations. A positive correlation between SLC25A32 dysfunction and flavoenzyme deficiency was observed. Besides the flavoenzymes involved in fatty acid ß-oxidation and amino acid metabolism being impaired, Slc25a32-/- embryos also had a subunit of glycine cleavage system-dihydrolipoamide dehydrogenase damaged, resulting in glycine accumulation and glycine derived-formate reduction, which further disturbed folate-mediated one-carbon metabolism, leading to 5-methyltetrahydrofolate shortage and other folate intermediates accumulation. Maternal formate supplementation increased the 5-methyltetrahydrofolate levels and ameliorated the NTDs in Slc25a32-/- embryos. The Slc25a32K235R/K235R and Slc25a32Y174C/K235R mice had no glycine accumulation, but had another formate donor-dimethylglycine accumulated and formate deficiency. Meanwhile, they suffered from the absence of all folate intermediates in mitochondria. Formate supplementation increased the folate amounts, but this effect was not restricted to the Slc25a32 mutant mice only. In summary, we established novel animal models, which enabled us to understand the function of SLC25A32 better and to elucidate the role of SLC25A32 dysfunction in human disease development and progression.

FMR1 allele frequencies in 51,000 newborns: a large-scale population study in China.

BACKGROUND: Fragile X syndrome (FXS), caused by CGG-repeat expansion in FMR1 promoter, is one of the most common causes of mental retardation. Individuals with full mutation and premutation alleles have a high risk of psychophysiological disorder and of having affected offspring. Frequencies of FMR1 alleles in general newborns have been reported in Caucasians but have not been investigated in the large-scale population in  the mainland of China. METHODS: The sizes of FMR1 CGG-repeats were analyzed in 51,661 newborns (28,114 males and 23,547 females) and also in a cohort of 33 children diagnosed with developmental delay using GC-rich polymerase chain reaction (PCR) and triple repeat primed PCR. RESULTS: The frequency of CGG repeats > 100 was 1/9371 in males and 1/5887 in females, and the frequency of CGG repeats > 54 was 1/1561 in males and 1/1624 in females. FMR1 full mutation and premutation were identified in 27.27% of children who had Ages and Stages Questionnaire scores less than two standard deviations from the cutoff value. CONCLUSIONS: Our study revealed the prevalence of FXS in China and improved the sample databases of FXS, suggesting that the prevalence of FXS in Chinese is higher than estimated previously and that FXS screening can be advised to high-risk families.

Constructing 24(23→22)-abeo-Cholestane from Tigogenin in a 20(22→23)-abeo-Way via a PhI(OAc)2-mediated Favorskii Rearrangement.

Transforming tigogenin, a steroidal sapogenin, to a 24(23→22)-abeo-cholestane, which is an unusual structural feature shared by the aglycons of saundersiosides and candicanoside A, is described. The spiroketal of tigogenin was unfolded and the resulting C22-ketone was subjected to Favorskii rearrangement mediated by PhI(OAc)2/KOH/MeOH to squeeze out the C22 from the side chain, thus reaching the 24(23→22)-abeo-cholestane structure.

Synthesis of 12,12'-azo-13,13'-diepi-Ritterazine N.

A synthesis of the 12,12'-azo-analogue of ritterazine N from hecogenin is reported. Ring contraction of two 6/5 bicyclic ring systems, one trans-fused and another spiro, to 5/5 spiro ring systems is accomplished with excellent stereochemical control. Key transformations include an abnormal Baeyer-Villiger oxidation, a Norrish type I cleavage, an intramolecular dipolar [3 + 2] cycloaddition, and an intramolecular oxymecuration. Failing to uncover the ß-OH ketone from the isoxazoline ring, we end up with a synthesis of a cyclic analogue of ritterazine N.

Complete sequence of a hepatic cirrhosis inbred C57BL/6 mice model mitochondrial genome.

In the present work we undertook the complete mitochondrial genome sequencing of an important hepatic cirrhosis model inbred C57BL/6 strain for the first time. Its mitogenome was 16,312 bp and coding 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes. A total of 96 SNPs were examined when compared to reference BN sequence.

Concise synthesis of the core structures of saundersiosides.

A divergent synthesis of three core pentacyclic lactones of nine rearranged cholestane sapogenins, saundersiosides A-H (1-8) and candicanoside A (9), is reported. Key features include a one-flask CBS reduction/Brown hydroboration-oxidation, a SmI2-mediated intramolecular Reformatskii reaction, and an intramolecular transesterification. This synthesis provides a general strategy and key precursors for the collective synthesis of natural and designed saundersiosides. An efficient formal synthesis of candicanoside A is also achieved.

A blunted sympathetic and accentuated parasympathetic response to postural change in subjects with depressive disorders.

BACKGROUND: In recent years, the bi-directional relationship between depression and ANS dysfunction has received considerable attention, but findings remain inconclusive. In this study, we aimed to examine the spectral HRV response to postural change in subjects with depressive disorders and in healthy controls, in order to gain insight into the characteristics of autonomic nervous system (ANS) response to postural change in subjects with depressive disorders. METHODS: We compared HRV response to postural change between subjects with depressive disorders and healthy controls aged 20-37 years. Depression severity was assessed by the self-reported Beck Depression Inventory-II (BDI-II). Spectral HRV was analyzed at two moments: 10 min seated rest and 10 min at standing position, with spontaneous breathing. RESULTS: No significant differences existed in the resting spectral HRV indices between subjects with depressive disorders and controls, however, following postural change, the increasing level of LF and LF/HF was lower and the decreasing level of HF power was higher, in the individuals with depression than that in healthy subjects. The differences in the LF power, HF power and the LF/HF ratio between seated rest before standing up and after postural change were found negatively correlated with depression severity. CONCLUSION: We found a blunted sympathetic and accentuated parasympathetic response to postural change in subjects with depressive disorder, suggesting that the autonomic impairment and early ANS dysfunction may exist among depressed individuals. These findings indicated that spectral analysis of HRV associated with postural change may be a more sensitive method than resting HRV analysis for detecting ANS dysfunction in depressive disorders. LIMITATIONS: Further studies are needed to expand the sample size and to clarify the mechanisms responsible for the autonomic dysfunction observed in individuals with depressive disorders.

Attenuated or absent HRV response to postural change in subjects with primary insomnia.

Previous studies have compared rest heart rate variability (HRV) between insomniacs and good sleepers, but the results have not been consistent. The altered HRV behavior in response to postural change was considered useful as another sensitive measure for evaluating the autonomic nervous function, however, to our knowledge, no study was found using HRV response to postural change in primary insomnia. Our study aimed to examine HRV response to postural change maneuver (PCM) in both primary insomniacs and controls between 22 and 39 years of age to gain insights into the characteristics of the autonomic nervous system (ANS) function in primary insomnia subjects. HRV was recorded for 5 min at seated rest, and then, the subjects quickly stood up from a seated position in up to 3s and remained standing for 15 min. HRV was recorded at the following times: seated rest and 0-5 min, 5-10 min and 10-15 min in the standing position. In primary insomnia subjects, attenuated or absent HRV response to postural change was identified, the increase in LF/HF ratio and the decrease in HF and SD1 from seated to standing were much slower than in the normal controls. In conclusion, this study provided evidence of the possible bi-directional relationship between insomnia and autonomic nervous system (ANS) function, which will move us closer to developing a new sensitive method for measuring autonomic impairment and early sympathetic damage in primary insomnia subjects.

Tanshinone IIA therapeutically reduces LPS-induced acute lung injury by inhibiting inflammation and apoptosis in mice.

AIM: To study the effects of tanshinone IIA (TIIA) on lipopolysaccharide (LPS)-induced acute lung injury in mice and the underlying mechanisms. METHODS: Mice were injected with LPS (10 mg/kg, i.p.), then treated with TIIA (10 mg/kg, i.p.). Seven hours after LPS injection, the lungs were collected for histological study. Protein, LDH, TNF-α and IL-1ß levels in bronchoalveolar lavage fluid (BALF) and myeloperoxidase (MPO) activity in lungs were measured. Cell apoptosis and Bcl-2, caspase-3, NF-κB and HIF-1α expression in lungs were assayed. RESULTS: LPS caused marked histological changes in lungs, accompanied by significantly increased lung W/D ratio, protein content and LDH level in BALF, and Evans blue leakage. LPS markedly increased neutrophil infiltration in lungs and inflammatory cytokines in BALF. Furthermore, LPS induced cell apoptosis in lungs, as evidenced by increased TUNEL-positive cells, decreased Bcl-2 content and increased cleaved caspase-3 content. Moreover, LPS significantly increased the expression of NF-κB and HIF-1α in lungs. Treatment of LPS-injected mice with TIIA significantly alleviated these pathological changes in lungs. CONCLUSION: TIIA alleviates LPS-induced acute lung injury in mice by suppressing inflammatory responses and apoptosis, which is mediated via inhibition of the NF-κB and HIF-1α pathways.

Lentiviral miR30-based RNA interference against heparanase suppresses melanoma metastasis with lower liver and lung toxicity.

AIM: To construct short hairpin RNAs (shRNAs) and miR30-based shRNAs against heparanase (HPSE) to compare their safety and their effects on HPSE down-modulation in vitro and in vivo to develop a more ideal therapeutic RNA interference (RNAi) vector targeting HPSE. METHODS: First, we constructed shRNAs and miR30-based shRNAs against HPSE (HPSE-shRNAs and HPSE-miRNAs) and packed them into lentiviral vectors. Next, we observed the effects of the shRNAs on knockdown for HPSE expression, adhesion, migration and invasion abilities in human malignant melanoma A375 cells in vitro. Furthermore, we compared the effects of the shRNAs on melanoma growth, metastasis and safety in xenograft models. RESULTS: Our data showed that these artificial miRNAs targeting HPSE could be effective RNAi agents mediated by Pol II promoters in vitro and in vivo, although these miRNAs were not more potent than the HPSE-shRNAs. It was noted that obvious lung injuries, rarely revealed previously, as well as hepatotoxicity could be caused by lentivirus-mediated shRNAs (LV shRNAs) rather than lentivirus-mediated miRNAs (LV miRNAs) in vivo. Furthermore, enhanced expression of pro-inflammatory cytokines IL-6 and TGF-ß1 and endogenous mmu-miR-21a-5p were detected in lung tissues of shRNAs groups, whereas the expression of mmu-let-7a-5p, mmu-let-7b-5p and mmu-let-7c-5p were down-regulated. CONCLUSION: These findings suggest that artificial miRNAs display an improved safety profile of lowered lung injury or hepatotoxicity relative to shRNAs in vivo. The mechanism of lung injuries caused by shRNAs may be correlated with changes of endogenous miRNAs in the lung. Our data here increase the flexibility of a miRNA-based RNAi system for functional genomic and gene therapy applications.

[Effects of harvest date on quality of BNS hybrid wheat flour and steamed bread].

Three wheat hybrids derived from the BNS male sterile line were used to study the effect of harvest time on gluten content, dough rheological properties, starch pasting properties of wheat flour and the quality of steamed bread. Each of the three hybrids was harvested at May 27, May 31 and June 4, respectively. The results showed that the flour quality of BNS hybrids was affected by harvest time to some extents, and the best harvest time varied among the different hybrids. For Baiza 1, Baiza 2 and Baiza 3, the best harvest time was May 27, May 27 and June 4, respectively. Both the flour quality and the comprehensive score and taste of steamed bread were the best for the hybrids harvested at these dates. Moreover, Baiza 2 was suited to make steamed bread and noodle when harvested on May 27.

Involvement of microRNA-mediated gene expression regulation in the pathological development of stem canker disease in Populus trichocarpa.

MicroRNAs (miRNAs), a type of short (21-23 nucleotides), non-coding RNA molecule, mediate repressive gene regulation through RNA silencing at the post-transcriptional level, and play an important role in defense and response to abiotic and biotic stresses. In the present study, Affymetrix® miRNA Array, real-time quantitative PCR (qPCR) for miRNAs and their targets, and miRNA promoter analysis were used to validate the gene expression patterns of miRNAs in Populus trichocarpa plantlets induced with the poplar stem canker pathogen, Botryosphaeria dothidea. Twelve miRNAs (miR156, miR159, miR160, miR164, miR166, miR168, miR172, miR319, miR398, miR408, miR1448, and miR1450) were upregulated in the stem bark of P. trichocarpa, but no downregulated miRNAs were found. Based on analysis of the miRNAs and their targets, a potential co-regulatory network was developed to describe post-transcriptional regulation in the pathological development of poplar stem canker. There was highly complex cross-talk between diverse miRNA pathway responses to biotic and abiotic stresses. The results suggest that miR156 is probably an integral component of the miRNA response to all environmental stresses in plants. Cis-regulatory elements were binding sites for the transcription factors (TFs) on DNA. Promoter analysis revealed that TC-rich repeats and a W1-box motif were both tightly related disease response motifs in Populus. Promoter analysis and target analysis of miRNAs also revealed that some TFs regulate their activation/repression. Furthermore, a feedback regulatory network in the pathological development of poplar stem canker is provided. The results confirm that miRNA pathways regulate gene expression during the pathological development of plant disease, and provide new insights into understanding the onset and development of poplar stem canker.

Case-control study on the association between qi-stagnation and insomnia.

OBJECTIVE: To investigate the relationship between insomnia and qi-stagnation by using the international standardized measurement of sleep quality and the Traditional Chinese Medicine (TCM) Constitution Scales. METHODS: A survey by means of the TCM Constitution Scales, the Pittsburgh Sleep Quality Index (PSQI), Epworth Sleepiness Scale (ESS) and the Deep Sleep Scale (DSS) in 169 participants aged between 16 and 80 years old was conducted. Comparison was made to examine the sleep quality and insomnia symptoms in the qi-stagnation group and other-constitution group. RESULTS: Univariate analysis found that the qi-stagnation group had a significantly increased risk of difficulty in falling asleep (OR=3.012, and 95% CI 1.310 to 6.923 for PSQI; OR=3.016, and 95% CI 1.358 to 6.709 for DSS) and early waking (OR=3.545, and 95% CI 1.229 to 10.232 for PSQI; OR=2.742, and 95% CI 1.072 to 7.014 for DSS), while the other-constitution group had a significant risk of dreaminess (OR=2.419, and 95% CI 1.154 to 5.072 for PSQI; OR=2.561, and 95% CI 1.116 to 5.880 for DSS). A dose-effect relationship existed between insomnia symptoms and qi-stagnation. Qi-stagnation significantly increased the risk of difficulty in falling asleep and early waking. CONCLUSION: This case-control study revealed that there is a statistically significant association between qi-stagnation and insomnia. Based on this study, we recommend that further research should be conducted for the rehabilitative care and cure of insomnia from the perspective of TCM constitution.

[Construction of VEGF gene-targeted small interfering RNA mediated by CMV promoter and its silencing effect].

OBJECTIVE: To construct VEGF gene-targeted small interfering RNA (siRNA) and its expression vector driven by CMV promoter and to investigate its interference effect. METHODS: The VEGF gene-targeted hairpin siRNA was designed, two complementary oligonucleotide strands were synthesized. After annealing, two-strand oligonucleotide was inserted into pDC311-SV40-RC vector, which was then identified by PCR and sequenced. Then human U-2 OS cell line was transfected with the vector using lipofectamine method. Finally, ELISA was performed to evaluate the expression of VEGF protein. RESULTS: PCR-identification of positive clone and sequencing confirmed the vector containing the target siRNA. ELISA showed that compared with the control group, the expression levels of VEGF protein in transfected U-2 OS cells were decreased significantly (P<0.05). CONCLUSION: VEGF gene-targeted siRNA and its vector mediated by CMV promoter were successfully constructed, which can reduce the VEGF protein expression after transfecting.

A genetic map constructed using a doubled haploid population derived from two elite Chinese common wheat varieties.

Genetic mapping provides a powerful tool for the analysis of quantitative trait loci (QTLs) at the genomic level. Herein, we report a new genetic linkage map developed from an F(1)-derived doubled haploid (DH) population of 168 lines, which was generated from the cross between two elite Chinese common wheat (Triticum aestivum L.) varieties, Huapei 3 and Yumai 57. The map contained 305 loci, represented by 283 simple sequence repeat (SSR) and 22 expressed sequence tag (EST)-SSR markers, which covered a total length of 2141.7 cM with an average distance of 7.02 cM between adjacent markers on the map. The chromosomal locations and map positions of 22 new SSR markers were determined, and were found to distribute on 14 linkage groups. Twenty SSR loci showed different chromosomal locations from those reported in other maps. Therefore, this map offers new information on the SSR markers of wheat. This genetic map provides new opportunities to detect and map QTLs controlling agronomically important traits. The unique features of this map are discussed.

Matrine inhibits invasiveness and metastasis of human malignant melanoma cell line A375 in vitro.

BACKGROUND: Matrine is a traditional Chinese medicine with significant inhibitory activity against malignant tumors. Its effects on the invasiveness and metastasis of malignant tumors have rarely been reported. AIM: To investigate whether matrine can inhibit the metastasis-related activities of the human malignant melanoma cell line A375 in vitro. METHODS: 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and Annexin-V-fluorescein isothiocyanate/propidium iodide (Annexin-V-FITC/PI) affinity assay were used to examine the effects of matrine on the proliferation and apoptosis induction of A375 cells. The morphologic changes of A375 cells were observed by light and electron microscopy. Semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting were performed to evaluate the expression of heparanase mRNA and protein. The effect of matrine on the adhesion ability and invasiveness of treated A375 cells was tested by cell-Matrigel adhesion assay and Matrigel invasion assay, respectively. RESULTS: Matrine showed significant inhibition of the proliferation of A375 cells in a dose- and time-dependent manner. It also induced apoptosis in a dose-dependent manner. Compared with the control group, the levels of heparanase mRNA and protein expression of A375 cells treated with different concentrations of matrine were decreased significantly, as were their adhesion ability and invasiveness. CONCLUSIONS: These findings indicate that matrine inhibits the invasiveness and metastasis of A375 cells in vitro. The mechanisms may be linked to the inhibition of cellular proliferation, induction of apoptosis, and downregulation of heparanase mRNA and protein expression.

Invariant natural killer T cells from rhesus macaque spleen and peripheral blood are phenotypically and functionally distinct populations.

BACKGROUND: Natural killer T cells (NKT) possess dual functions of innate and adaptive immune systems, controlling viral infections and regulating autoimmune diseases. Non-human primates (NHP) are penultimate models for advancing therapeutic immunoregulatory strategies for translational application in humans, though, little is known about NHP NKT cells. Here we characterized rhesus macaque NKT cells ex vivo. METHODS: The frequency, phenotype and intracellular cytokine production of V alpha 24+ 6B11+ invariant NKT (iNKT) cells were analyzed by multi-color flow cytometry. V alpha 24J alpha Q mRNA expression was analyzed by real-time RT-PCR. RESULTS: The frequencies of peripheral blood (PB) and spleen V alpha 24+ 6B11+ iNKT cells were not significantly different. The iNKT cell subset in spleen was significantly increased for CD4+ CD8+ and CD3+ CD56+ co-expression as well as intracellular interleukin-4 production, which was rarely observed in circulating PB. CONCLUSION: Spleen iNKT cells in rhesus macaques are Th2 biased and display phenotypically and functionally distinct profiles from their PB counterpart.