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Abstract Background Dendrobium catenatum is a perennial herb of the genus Dendrobium orchidaceae. It has been known as “Golden Grass, Soft Gold” since ancient times with effects of strengthening the body, benefiting the stomach, generating body fluid, nourishing Yin and clearing internal heat. The flowers of D. catenatum have anti-oxidation, immune regulation and other biological activities. The composition analysis of flowers showed that flavonoid glycosides were significantly accumulated in floral tissue. However, in the flowers of D. catenatum, there was only one case of the UDP-glycosyltransferase (UGT) responsible for the glycosylation of flavonoids has been reported. Result In this study, a new UGT (named UGT708S6) was cloned from D. catenatum flowers rich in O-glycosides and C-glycosides, and its function and biochemical properties were characterized. Through homology comparison and molecular docking, we identified the key amino acid residues affecting the catalytic function of UGT708S6. The glycosyltransferase UGT708S6 was characterized and demonstrated C-glycosyltransferase (CGT) activity in vitro assay using phloretin and 2-hydroxynaringenin as sugar acceptors. The catalytic promiscuity assay revealed that UGT708S6 has a clear sugar donor preference, and displayed O-glycosyltransferase (OGT) activity towards luteolin, naringenin and liquiritigenin. Furthermore, the catalytic characteristics of UGT708S6 were explored, shedding light on the structural basis of substrate promiscuity and the catalytic mechanism involved in the formation of flavonoid C-glycosides. R271 was a key amino acid residue site that sustained the catalytic reaction. The smaller binding pocket resulted in the production of new O-glycosides and the reduction of C-glycosides. This highlighted the importance of the binding pocket in determining whether C-glycosides or O-glycosides were produced. Conclusions The findings suggest that UGT708S6 holds promise as a new glycosyltransferase for synthesizing flavonoid glycosides and offer valuable insights for further understanding the catalytic mechanisms of flavonoid glycosyltransferases.

Abstract Amino acid/auxin permease (AAAP) genes encode a large family of protein transporters that play important roles in various aspects of plant growth and development. Here, we performed genome-wide identification of members in the foxtail millet (Setaria italica L.) AAAP family (SiAAAP) and their saline-alkali stress-induced expression patterns, resulting in the identification of 65 SiAAAP genes, which could be divided into eight subfamilies. Except for SiAAAP65, the remaining 64 genes were located on nine chromosomes of foxtail millet. Gene structure and conserved motif analyses indicated that the members in the same subfamily are highly conserved. Gene duplication event analysis suggested that tandem duplication may be the main factor driving the expansion of this gene family, and Ka/Ks analysis indicated that all the duplicated genes have undergone purifying selection. Transcriptome analysis showed differential expression of SiAAAPs in roots, stems, leaves, and tassel inflorescence. Analysis of cis-acting elements in the promoter indicated that SiAAAPs contain stress-responsive cis-acting elements. Under saline-alkali stress, qRT-PCR analysis showed that SiAAP3, SiLHT2, and SiAAP16 were differentially expressed between salt-alkali tolerant millet variety JK3 and salt-alkali sensitive millet variety B175. These results suggest that these genes may be involved in or regulate the response to saline-alkali stress, providing a theoretical basis for further studying the function of SiAAAPs.

AbstractContext Diabetic nephropathy (DN) is the main cause of end-stage renal disease. Modified Shen-Yan-Fang-Shuai formula (M-SYFSF) has excellent clinical efficacy in treating diabetic kidney disease. However, the potential mechanism of M-SYFSF remains unknown.Objective To investigate the mechanism of M-SYFSF against DN by network pharmacological analysis and biological experiments.Materials and methods Utilizing a web-based pharmacology database, the potential mechanisms of M-SYFSF against DN were identified. In vivo experiments, male SD rats were injected with streptozotocin (50 mg/kg) and got uninephrectomy to construct a model of DN. M-SYFSF (11.34 g/kg/d) was gavaged once per day for 12 weeks after model establishment. In vitro experiments, human proximal tubular cells (HK-2) were performed with advanced glycation end-products (AGEs) (100 μg/mL), then intervened with M-SYFSF freeze-dried powder. Pathological staining, WB, IHC, ELISA were conducted to explore the mechanism of M-SYFSF against DN.Results Network pharmacological analysis showed that MAPK pathway was the potential pathway. Results showed that compared with the Model group, M-SYFSF significantly reduced 24h urine albumin, UACR, and serum creatinine levels (54.90 ± 26.67 vs. 111.78 ± 4.28, 8.87 ± 1.69 vs. 53.94 ± 16.01, 11.56 ± 1.70 vs. 118.70 ± 49.57, respectively), and improved renal pathological changes. Furthermore, the intervention of M-SYFSF reduced the expression of pro-inflammatory cytokines and inhibited the activation of MAPK pathway in AGEs-treated HK-2 cells.Discussion and conclusion M-SYFSF is likely to reduce inflammation in DN by inhibiting the MAPK pathway. It provides a theoretical basis for the clinical application of M-SYFSF in the treatment of DN.

E3 ubiquitin ligases are central modifiers of plant signaling pathways that regulate protein function, localization, degradation, and other biological processes by linking ubiquitin to target proteins. E3 ubiquitin ligases include proteins with the U-box domain. However, there has been no report about the foxtail millet (Setaria italica L. Beauv) U-box gene family (SiPUB) to date. To explore the function of SiPUBs, this study performed genome-wide identification of SiPUBs and expression analysis of them in response to saline-alkali stress. A total of 70 SiPUBs were identified, which were unevenly distributed on eight chromosomes. Phylogenetic and conserved motif analysis demonstrated that SiPUBs could be clustered into six subfamilies (I–VI), and most SiPUBs were closely related to the homologues in rice. Twenty-eight types of cis-acting elements were identified in SiPUBs, most of which contained many light-responsive elements and plant hormone-responsive elements. Foxtail millet had 19, 78, 85, 18, and 89 collinear U-box gene pairs with Arabidopsis, rice, sorghum, tomato, and maize, respectively. Tissue specific expression analysis revealed great variations in SiPUB expression among different tissues, and most SiPUBs were relatively highly expressed in roots, indicating that SiPUBs may play important roles in root development or other growth and development processes of foxtail millet. Furthermore, the responses of 15 SiPUBs to saline-alkali stress were detected by qRT-PCR. The results showed that saline-alkali stress led to significantly differential expression of these 15 SiPUBs, and SiPUB20/48/70 may play important roles in the response mechanism against saline-alkali stress. Overall, this study provides important information for further exploration of the biological function of U-box genes.

The hepatitis C virus (HCV) causes severe liver diseases, including hepatitis, liver cirrhosis, and hepatocellular carcinoma, which have high morbidity and mortality. Antibody targeting receptor-mediated HCV infections have limited therapeutic benefits, suggesting that the transmission of HCV infections is possibly mediated via receptor-independent mechanisms. Exosomes are membrane-enclosed vesicles with a diameter of 30–200 nm, which originate from the fusion of endosomal multivesicular bodies with the plasma membrane. Accumulating evidence suggests that exosomes have a pivotal role in HCV infections. Exosomes can transfer viral and cellular bioactive substances, including nucleic acids and proteins, to uninfected cells, thus spreading the infection by masking these materials from immunological recognition. In addition, exosomes originating from some cells can deliver antiviral molecules or prompt the immune response to inhibit HCV infection. Exosomes can be used for the diagnosis of HCV-related diseases, and are being presently evaluated as therapeutic tools for anti-HCV drug delivery. This review summarizes the current knowledge on the dual roles and potential clinical applications of exosomes in HCV infections.

Alkaline/neutral invertase (NI) proteins irreversibly break down sucrose into glucose and fructose and play a vital role in carbohydrate and plant growth and development. Currently, a comprehensive understanding of NI genes in Dendrobium catenatum is lacking. After genome-wide identification and prediction, nine DcNI genes were identified and named DcNI1–DcNI9. Through comprehensive analysis of gene structure, amino acid sequence, conserved motif and phylogenetic tree, it was found that the DcNI gene family has a typical conserved region and its amino acid sequence is similar to other plant NI genes. However, DcNI3 lost amino acid sequences at both the N-terminus and C-terminus, while DcNI8 lost amino acids at the C-terminus. Phylogenetic analysis revealed that the DcNI family could be divided into α and β groups. It reveals conservation and divergence of DcNIs. Predictive analysis of cis-acting elements in the promoter region reveals the potential response of DcNI protein to hormones and environmental stress and suggests it to be widely involved in growth and development. DcNIs were differentially expressed in different tissues such as roots, stems and leaves. In particular, DcNI4 was highly expressed in the stem. They also had a differential expression response under drought, cold, P1 fungus infection and jasmonic acid (JA) treatment. To the best of our knowledge, this is the first genome-wide analysis of the DcNI gene family in D. catenatum. This study will provide important and primary information for the functional study of the DcNI gene family and the polysaccharide synthesis pathway in D. catenatum.

Dendrobium catenatum, which grows on the semi humid rocks in the mountains, has been at the top of the ‘Nine Immortals of China’ since ancient times. It is a kind of yin tonic medicine and its main active component is polysaccharide. Cellulose synthase-like D(CslD) genes were predicted to catalyse the biosynthesis of 1,4-β-d-glycan backbone of hemicelluloses, which plays fundamental roles in plant development. To investigate the role of CslD in the development of D. catenatum, eight CslD genes (DcCslD1,2a,2b,3a,3b,4a,4b,5) were identified. The results of protein prediction and analysis showed that CslD2a/2b/4a/4b proteins were acidic proteins. All the proteins had obvious hydrophobic or hydrophilic regions, and had transmembrane structure. Phylogenetic analysis revealed that the DcCslD family could be divided into group I, II, III and IV. DcCslD proteins had a typical Cellulose synthase domain and similar protein structures to the CslDs of other plants. Their promoter regions contain cis regulatory elements related to stress and hormone response. The results of qRT-PCR showed that the identified DcCslDs were differentially expressed in roots, stems and leaves. Most of them were highly expressed in stems and leaves. The environmental stresses examination showed that the expression levels of DcCslD5 were closely associated with drought-recovery treatment; the expression levels of DcCslD1, DcCslD2a, DcCslD2b, DcCslD3a and DcCslD5 were significantly influenced by low temperature. This study systematically analyzed the sequence characteristics of CslD protein of D. catenatum, which can provide reference for further study on the function of CslD protein in the polysaccharide metabolism of D. catenatum. Supplemental data for this article is available online at https://doi.org/10.1080/13102818.2021.1941252 .

Anthracnose, caused by Colletotrichum gloeosporioides, is highly harmful for walnut production in the world. To better control this disease, the inhibitory effects of 3 fungicides against Colletotrichum gloeosporioides were determined, and the results showed that prochloraz had better inhibitory activity. Through comparative metabolomics analysis, 311 metabolites might be associated with the walnut peel response to Colletotrichum gloeosporioides under prochloraz treatment. Furthermore, we supposed that the phenylpropanoid pathway might be induced by prochloraz to resist Colletotrichum gloeosporioides infection. In conclusion, the upregulated metabolites in the phenylpropanoid pathway might be related to synthesize lignin to further form a cell wall against Colletotrichum gloeosporioides infection.

It has been reported that autophagic stress, which is involved in many diseases, plays a key role in the development of diabetic nephropathy (DN). In this study, we investigated the effects of high dose vitamin E on renal tubular epithelial cells and autophagic stress-related mechanisms in diabetes condition. In diabetic rats, high dose vitamin E treatment significantly decreased the serum creatinine, urea nitrogen, urinary albumin and urinary protein, reduced the levels of LCN2, HAVCR1, LDH and 8-OHdG in urine, and attenuated the cellular apoptosis and interstitial fibrosis in renal cortex. In vitro, vitamin E could reduce the release of LCN2 and HAVCR1 and the protein levels of caspase 3 and TGF-β1, as well as improve the growth inhibition in cultured HK-2 cells after exposure to advanced glycation end products (AGEs). Also, LC3-II and SQSTM1-positive dots were significantly increased in the renal tubular epithelial cells of DN patients and diabetic rats, and in HK-2 cells after exposure to AGEs, which were markedly declined by vitamin E. In addition, we found that the autophagosome formation was not affected by AGEs, as assessed by the mRNA levels of LC3B, Beclin-1, and ATG7. However, AGEs blocked the lysosomal degradation of autophagosome, which was characterized by a decrease in the enzymatic activity of cathepsin B/cathepsin L and DQ-ovalbumin degradation in HK-2 cells, indicating that AGEs-induced accumulation of autophagic vacuoles was a sign of autophagic stress. Interestingly, vitamin E exerted a protective effect on lysosomes to reduce the autophagic stress. Taken together, we conclude that autophagic stress may play an important part in the progression of DN, and alleviation of autophagic stress though improvement of lysosomal function provides a promising novel approach for treating DN.

Acupuncture at homotopic acupoints or heterotopic acupoints is known to either inhibit or facilitate gastrointestinal motility, depending on the acupoint location. However, little effort has been made to investigate the roles of specific receptors (such as adrenergic and muscarinic acetylcholine receptors) in mediating the effects of acupuncture at heterotopic and homotopic acupoints. Different adrenergic receptor subtypes or cholinergic receptor subtypes are predominantly expressed in various sections of the gut, resulting in variations between the effects of acupuncture at heterotopic or homotopic acupoints on gastrointestinal motility. Here, we investigated the role of β1/β2 receptors and M2/M3 receptors in gastrointestinal motility regulated by acupuncture at ST37, a heterotopic acupoint, and ST25, a homotopic acupoint, by simultaneously recording intraluminal pressures in the distal colon and stomach or jejunum and examining fecal phenol red excretion in β1/2 receptor-knockout mice and M2/3 receptor-knockout mice. We found that knockout of the M2/3 receptor significantly inhibited ST37 acupuncture-induced enhancement of gastric motility, jejunal motility, and colonic motility. Additionally, knocking out of the β1/2 receptor significantly diminished the ST25 acupuncture-induced inhibition of gastric motility and jejunal motility without significantly altering the enhancement of colonic motility induced by acupuncture at ST25. Acupuncture at ST37 significantly accelerated gastrointestinal transition in β1/2 receptor-knockout mice and their wild-type littermates. However, this acceleration of gastrointestinal transition was markedly diminished in M2/3 receptor-knockout mice relative to their wild-type littermates. Acupuncture at ST25 significantly increased gastrointestinal transition in β1/2 receptor-knockout mice and significantly decreased gastrointestinal transition in M2/3 receptor-knockout mice without altering gastrointestinal transition in wild-type littermates of either. Our study revealed that M2/3 receptors are required for the gastrointestinal motility associated with whole gastrointestinal transition enhanced by acupuncture at heterotopic acupoints, whereas β1/2 receptors are required for the same gastrointestinal motility processes inhibited by acupuncture at homotopic acupoints. Therefore, our findings reveal important biological mechanisms underlying acupuncture treatment of disorders involving gastrointestinal motility dysfunction.

Background: The previous study showed that the cardiac arrhythmias induced by myocardial ischemia and reperfusion were attenuated by the pretreatment of acupuncture; however, the related mechanism is not understood. The present study was therefore designed to determine whether intracellular Ca2+ ([Ca2+]i) and connexin 43 (Cx43) are involved in the mediation of the anti-arrhythmic effect of electro-acupuncture (EA) pretreatment in the rats subjected to simulative global ischemia and reperfusion (SGIR). Methods: SGIR was made in the isolated heart by a low flow perfusion followed by a flow restoration. Four groups of animals are involved in the present study, including normal control group, SGIR group, EA group and EA plus 18 beta-glycyrrhetinic acid (EAG) group. For EA pretreatment, bilateral Neiguan acupoints (PC6) of the rats were stimulated for 30 min once a day in 3 consecutive days. Cx43 antagonist was given to the rats in EAG group 30 minutes before the EA pretreatment. The resting [Ca2+]i concentration, calcium oscillation, the contents of total Cx43 and non-phosphrylated Cx43 and arrhythmia score were compared among different groups. Results: In EA group, the arrhythmic score, the resting [Ca2+]i concentration and the number of [Ca2+]i oscillations were all significantly less than those in SGIR group (all P < 0.05), and interestingly, after EA pretreatment, the contents of nonphosphated Cx43 in the EA group were significantly lower than that in SGIR group respectively (P < 0.05). However, when the rats were treated with Cx43 antagonist prior to the EA pretreatment, the protection effects induced by EA pretreatment were reversed. Conclusions: The results showed that EA pretreatment could produce anti-arrhythmic effect in the rats subjected to SGIR. The anti-arrhythmic effect of EA pretreatment may be due at least partially to the inhibition of SGIR-induced calcium overload and [Ca2+]i oscillations, reduction of non-phosphorylated Cx43 and the enhancement of the corresponding phosphorylated Cx43 in the cardiac cells. [ABSTRACT FROM AUTHOR]

Background: Dendrobium catenatum is a perennial herb of the genus Dendrobium orchidaceae. It has been known as "Golden Grass, Soft Gold" since ancient times with effects of strengthening the body, benefiting the stomach, generating body fluid, nourishing Yin and clearing internal heat. The flowers of D. catenatum have anti-oxidation, immune regulation and other biological activities. The composition analysis of flowers showed that flavonoid glycosides were significantly accumulated in floral tissue. However, in the flowers of D. catenatum, there was only one case of the UDP-glycosyltransferase (UGT) responsible for the glycosylation of flavonoids has been reported. Result: In this study, a new UGT (named UGT708S6) was cloned from D. catenatum flowers rich in O-glycosides and C-glycosides, and its function and biochemical properties were characterized. Through homology comparison and molecular docking, we identified the key amino acid residues affecting the catalytic function of UGT708S6. The glycosyltransferase UGT708S6 was characterized and demonstrated C-glycosyltransferase (CGT) activity in vitro assay using phloretin and 2-hydroxynaringenin as sugar acceptors. The catalytic promiscuity assay revealed that UGT708S6 has a clear sugar donor preference, and displayed O-glycosyltransferase (OGT) activity towards luteolin, naringenin and liquiritigenin. Furthermore, the catalytic characteristics of UGT708S6 were explored, shedding light on the structural basis of substrate promiscuity and the catalytic mechanism involved in the formation of flavonoid C-glycosides. R271 was a key amino acid residue site that sustained the catalytic reaction. The smaller binding pocket resulted in the production of new O-glycosides and the reduction of C-glycosides. This highlighted the importance of the binding pocket in determining whether C-glycosides or O-glycosides were produced. Conclusions: The findings suggest that UGT708S6 holds promise as a new glycosyltransferase for synthesizing flavonoid glycosides and offer valuable insights for further understanding the catalytic mechanisms of flavonoid glycosyltransferases. [ABSTRACT FROM AUTHOR]

Diabetic nephropathy (DN) is the main cause of end-stage renal disease. Modified Shen-Yan-Fang-Shuai formula (M-SYFSF) has excellent clinical efficacy in treating diabetic kidney disease. However, the potential mechanism of M-SYFSF remains unknown. To investigate the mechanism of M-SYFSF against DN by network pharmacological analysis and biological experiments. Utilizing a web-based pharmacology database, the potential mechanisms of M-SYFSF against DN were identified. In vivo experiments, male SD rats were injected with streptozotocin (50 mg/kg) and got uninephrectomy to construct a model of DN. M-SYFSF (11.34 g/kg/d) was gavaged once per day for 12 weeks after model establishment. In vitro experiments, human proximal tubular cells (HK-2) were performed with advanced glycation end-products (AGEs) (100 μg/mL), then intervened with M-SYFSF freeze-dried powder. Pathological staining, WB, IHC, ELISA were conducted to explore the mechanism of M-SYFSF against DN. Network pharmacological analysis showed that MAPK pathway was the potential pathway. Results showed that compared with the Model group, M-SYFSF significantly reduced 24h urine albumin, UACR, and serum creatinine levels (54.90 ± 26.67 vs. 111.78 ± 4.28, 8.87 ± 1.69 vs. 53.94 ± 16.01, 11.56 ± 1.70 vs. 118.70 ± 49.57, respectively), and improved renal pathological changes. Furthermore, the intervention of M-SYFSF reduced the expression of pro-inflammatory cytokines and inhibited the activation of MAPK pathway in AGEs-treated HK-2 cells. M-SYFSF is likely to reduce inflammation in DN by inhibiting the MAPK pathway. It provides a theoretical basis for the clinical application of M-SYFSF in the treatment of DN. [ABSTRACT FROM AUTHOR]

Anthracnose, caused by Colletotrichum gloeosporioides, is highly harmful for walnut production in the world. To better control this disease, the inhibitory effects of 3 fungicides against Colletotrichum gloeosporioides were determined, and the results showed that prochloraz had better inhibitory activity. Through comparative metabolomics analysis, 311 metabolites might be associated with the walnut peel response to Colletotrichum gloeosporioides under prochloraz treatment. Furthermore, we supposed that the phenylpropanoid pathway might be induced by prochloraz to resist Colletotrichum gloeosporioides infection. In conclusion, the upregulated metabolites in the phenylpropanoid pathway might be related to synthesize lignin to further form a cell wall against Colletotrichum gloeosporioides infection. [ABSTRACT FROM AUTHOR]

Dendrobium catenatum, which grows on the semi humid rocks in the mountains, has been at the top of the 'Nine Immortals of China' since ancient times. It is a kind of yin tonic medicine and its main active component is polysaccharide. Cellulose synthase-like D(CslD) genes were predicted to catalyse the biosynthesis of 1,4-β-d-glycan backbone of hemicelluloses, which plays fundamental roles in plant development. To investigate the role of CslD in the development of D. catenatum, eight CslD genes (DcCslD1,2a,2b,3a,3b,4a,4b,5) were identified. The results of protein prediction and analysis showed that CslD2a/2b/4a/4b proteins were acidic proteins. All the proteins had obvious hydrophobic or hydrophilic regions, and had transmembrane structure. Phylogenetic analysis revealed that the DcCslD family could be divided into group I, II, III and IV. DcCslD proteins had a typical Cellulose synthase domain and similar protein structures to the CslDs of other plants. Their promoter regions contain cis regulatory elements related to stress and hormone response. The results of qRT-PCR showed that the identified DcCslDs were differentially expressed in roots, stems and leaves. Most of them were highly expressed in stems and leaves. The environmental stresses examination showed that the expression levels of DcCslD5 were closely associated with drought-recovery treatment; the expression levels of DcCslD1, DcCslD2a, DcCslD2b, DcCslD3a and DcCslD5 were significantly influenced by low temperature. This study systematically analyzed the sequence characteristics of CslD protein of D. catenatum, which can provide reference for further study on the function of CslD protein in the polysaccharide metabolism of D. catenatum. Supplemental data for this article is available online at https://doi.org/10.1080/13102818.2021.1941252. [ABSTRACT FROM AUTHOR]

Alkaline/neutral invertase (NI) proteins irreversibly break down sucrose into glucose and fructose and play a vital role in carbohydrate and plant growth and development. Currently, a comprehensive understanding of NI genes in Dendrobium catenatum is lacking. After genome-wide identification and prediction, nine DcNI genes were identified and named DcNI1–DcNI9. Through comprehensive analysis of gene structure, amino acid sequence, conserved motif and phylogenetic tree, it was found that the DcNI gene family has a typical conserved region and its amino acid sequence is similar to other plant NI genes. However, DcNI3 lost amino acid sequences at both the N-terminus and C-terminus, while DcNI8 lost amino acids at the C-terminus. Phylogenetic analysis revealed that the DcNI family could be divided into α and β groups. It reveals conservation and divergence of DcNIs. Predictive analysis of cis-acting elements in the promoter region reveals the potential response of DcNI protein to hormones and environmental stress and suggests it to be widely involved in growth and development. DcNIs were differentially expressed in different tissues such as roots, stems and leaves. In particular, DcNI4 was highly expressed in the stem. They also had a differential expression response under drought, cold, P1 fungus infection and jasmonic acid (JA) treatment. To the best of our knowledge, this is the first genome-wide analysis of the DcNI gene family in D. catenatum. This study will provide important and primary information for the functional study of the DcNI gene family and the polysaccharide synthesis pathway in D. catenatum. [ABSTRACT FROM AUTHOR]

It has been reported that autophagic stress, which is involved in many diseases, plays a key role in the development of diabetic nephropathy (DN). In this study, we investigated the effects of high dose vitamin E on renal tubular epithelial cells and autophagic stress-related mechanisms in diabetes condition. In diabetic rats, high dose vitamin E treatment significantly decreased the serum creatinine, urea nitrogen, urinary albumin and urinary protein, reduced the levels of LCN2, HAVCR1, LDH and 8-OHdG in urine, and attenuated the cellular apoptosis and interstitial fibrosis in renal cortex. In vitro , vitamin E could reduce the release of LCN2 and HAVCR1 and the protein levels of caspase 3 and TGF-β1, as well as improve the growth inhibition in cultured HK-2 cells after exposure to advanced glycation end products (AGEs). Also, LC3-II and SQSTM1-positive dots were significantly increased in the renal tubular epithelial cells of DN patients and diabetic rats, and in HK-2 cells after exposure to AGEs, which were markedly declined by vitamin E. In addition, we found that the autophagosome formation was not affected by AGEs, as assessed by the mRNA levels of LC3B, Beclin-1, and ATG7. However, AGEs blocked the lysosomal degradation of autophagosome, which was characterized by a decrease in the enzymatic activity of cathepsin B/cathepsin L and DQ-ovalbumin degradation in HK-2 cells, indicating that AGEs-induced accumulation of autophagic vacuoles was a sign of autophagic stress. Interestingly, vitamin E exerted a protective effect on lysosomes to reduce the autophagic stress. Taken together, we conclude that autophagic stress may play an important part in the progression of DN, and alleviation of autophagic stress though improvement of lysosomal function provides a promising novel approach for treating DN. [ABSTRACT FROM AUTHOR]

Acupuncture at homotopic acupoints or heterotopic acupoints is known to either inhibit or facilitate gastrointestinal motility, depending on the acupoint location. However, little effort has been made to investigate the roles of specific receptors (such as adrenergic and muscarinic acetylcholine receptors) in mediating the effects of acupuncture at heterotopic and homotopic acupoints. Different adrenergic receptor subtypes or cholinergic receptor subtypes are predominantly expressed in various sections of the gut, resulting in variations between the effects of acupuncture at heterotopic or homotopic acupoints on gastrointestinal motility. Here, we investigated the role of β1/β2 receptors and M2/M3 receptors in gastrointestinal motility regulated by acupuncture at ST37, a heterotopic acupoint, and ST25, a homotopic acupoint, by simultaneously recording intraluminal pressures in the distal colon and stomach or jejunum and examining fecal phenol red excretion in β1/2 receptor-knockout mice and M2/3 receptor-knockout mice. We found that knockout of the M2/3 receptor significantly inhibited ST37 acupuncture-induced enhancement of gastric motility, jejunal motility, and colonic motility. Additionally, knocking out of the β1/2 receptor significantly diminished the ST25 acupuncture-induced inhibition of gastric motility and jejunal motility without significantly altering the enhancement of colonic motility induced by acupuncture at ST25. Acupuncture at ST37 significantly accelerated gastrointestinal transition in β1/2 receptor-knockout mice and their wild-type littermates. However, this acceleration of gastrointestinal transition was markedly diminished in M2/3 receptor-knockout mice relative to their wild-type littermates. Acupuncture at ST25 significantly increased gastrointestinal transition in β1/2 receptor-knockout mice and significantly decreased gastrointestinal transition in M2/3 receptor-knockout mice without altering gastrointestinal transition in wild-type littermates of either. Our study revealed that M2/3 receptors are required for the gastrointestinal motility associated with whole gastrointestinal transition enhanced by acupuncture at heterotopic acupoints, whereas β1/2 receptors are required for the same gastrointestinal motility processes inhibited by acupuncture at homotopic acupoints. Therefore, our findings reveal important biological mechanisms underlying acupuncture treatment of disorders involving gastrointestinal motility dysfunction. [ABSTRACT FROM AUTHOR]