Study on the difference and correlation between the contents and toxicity of aristolochic acid analogues in Aristolochia plants.

ETHNOPHARMACOLOGICAL RELEVANCE: The nephrotoxicity and carcinogenicity induced by traditional Chinese medicines (TCMs) containing aristolochic acids (AAs) and related compound preparations have greatly limited their clinical application. While the toxicity of AA-I and AA-II is relatively clear, there are marked differences in the toxic effects of different types of aristolochic acid analogues (AAAs). Thus, the toxicity of TCMs containing AAAs cannot be evaluated based on the toxicity of a single compound. AIM OF THE STUDY: To systematically investigate the toxicity induced by Zhushalian (ZSL), Madouling (MDL) and Tianxianteng (TXT) as representative TCMs derived from Aristolochia. MATERIALS AND METHODS: AAA contents in ZSL, MDL and TXT were determined using HPLC. Subsequently, mice were treated for 2 weeks with high (H) and low (L) dosages of TCMs containing total AAA contents of 3 mg/kg and 1.5 mg/kg, respectively. Toxicity was evaluated using biochemical and pathological examination and was based on organ indices. Correlations between AAA contents and induced toxicity were analysed using multiple methods. RESULTS: Of the total AAA content, ZSL contained mainly AA-I and AA-II (>90%, of which AA-I accounted for 49.55%). AA-I accounted for 35.45% in MDL. TXT mainly contained AA-IVa (76.84%) and other AAAs accounted for <10%. Short-term toxicity tests indicated that ZSL and high-dose MDL induced obvious renal interstitial fibrosis and gastric injury, whereas TXT (high and low dosages) caused only slight toxicity. Correlation analysis suggested that AA-I might be the critical hazard factor for toxicity. CONCLUSIONS: The toxicity of TCMs containing AAAs cannot be generalised. The toxicity of TXT is relatively low compared with those of ZSL and MDL. The toxicity of Aristolochia depends mainly on the AA-I content; therefore, control of AA-I levels in TCMs and related compound preparations is required to reduce the risk of toxicity associated with the use of Aristolochia herbs in clinical settings.

Serum Amino Acids Imbalance in Canine Chronic Hepatitis: Results in 16 Dogs.

In humans, chronic liver disease may cause alterations in amino acids (AAs) metabolism, with serum branched-chain AAs (BCAAs) decreasing and aromatic AAs (AAAs) increasing. A reduced Fischer ratio (BCAAs/AAAs) has been found to be associated with hepatic fibrosis and is useful for assessing prognosis in human patients. In veterinary medicine, few studies have been performed, and in contrast to human patients, dogs with different kinds of hepatopathy tend to show both increased AAAs and BCAAs. In dogs, the association between histological scores and serum AAs has not been previously investigated. The aim of this study was to evaluate serum AAs in dogs with chronic hepatitis (CH) compared with a healthy control group (C) and, among CH dogs, in relation to their histological fibrosis and necroinflammatory activity scores. Leftover serum samples of 16 dogs with histological CH and 25 healthy dogs were employed. Serum AAs were measured by high performance liquid chromatography. Proline and the AAAs phenylalaine and tyrosine progressively increased with the histological severity. In contrast, cysteine, tryptophan and BCAA isoleucine progressively reduced. Lysine and the BCAAs leucine and valine showed a non-linear trend with the histological findings. The BCAAs/AAAs ratio was significatively reduced if higher grades of liver fibrosis were present.

Adult Mesenchymal Stem Cells and Derivatives in Improved Elastin Homeostasis in a Rat Model of Abdominal Aortic Aneurysms.

Abdominal aortic aneurysms (AAAs) are localized rupture-prone expansions of the aorta with limited reversibility that develop due to proteolysis of the elastic matrix. Natural regenerative repair of an elastic matrix is difficult due to the intrinsically poor elastogenicity of adult vascular smooth muscle cells (VSMCs). This justifies the need to provide external, pro-elastin regenerative- and anti-proteolytic stimuli to VSMCs in the AAA wall towards reinstating matrix structure in the aorta wall. Introducing alternative phenotypes of highly elastogenic and contractile cells into the AAA wall capable of providing such cues, proffers attractive prospects for AAA treatment. In this regard, we have previously demonstrated the superior elastogenicity of bone marrow mesenchymal stem cell (BM-MSC)-derived SMCs (cBM-SMCs) and their ability to provide pro-elastogenic and anti-proteolytic stimuli to aneurysmal SMCs in vitro. However, the major issues associated with cell therapy, such as their natural ability to home into the AAA tissue, their in vivo biodistribution and retention in the AAA wall, and possible paracrine effects on AAA tissue repair processes in the event of localization in remote tissues remain uncertain. Therefore, in this study we focused on assessing the fate, safety, and AAA reparative effects of BM-MSC-derived cBM-SMCs in vivo. Our results indicate that the cBM-SMCs (a) possess natural homing abilities similar to the undifferentiated BM-MSCs, (b) exhibit higher retention upon localization in the aneurysmal aorta than BM-MSCs, (c) downregulate the expression of several inflammatory and pro-apoptotic cytokines that are upregulated in the AAA wall contributing to accelerated elastic matrix breakdown and suppression of elastic fiber neo-assembly, repair, and crosslinking, and (d) improve elastic matrix content and structure in the AAA wall toward slowing the growth of AAAs. Our study provides initial evidence of the in vivo elastic matrix reparative benefits of cBM-SMCs and their utility in cell therapy to reverse the pathophysiology of AAAs.

Circular RNA Cdyl promotes abdominal aortic aneurysm formation by inducing M1 macrophage polarization and M1-type inflammation.

Macrophage polarization plays a crucial role in regulating abdominal aortic aneurysm (AAA) formation. Circular RNAs (circRNAs) are important regulators of macrophage polarization during the development of cardiovascular diseases. How-ever, the roles of circRNAs in regulating AAA formation through modulation of macrophage polarization remain unknown. In the present study, we compared circRNA microarray data under two distinct polarizing conditions (M1 and M2 macrophages) and identified an M1-enriched circRNA, circCdyl. Loss- and gain-of-function assay results demonstrated that circCdyl overexpression accelerated angiotensin II (Ang II)- and calcium chloride (CaCl2)-induced AAA formation by promoting M1 polarization and M1-type inflammation, while circCdyl deficiency showed the opposite effects. RNA pulldown, mass spectrometry analysis, and RNA immunoprecipitation (RIP) assays were conducted to elucidate the underlying mechanisms by which circCdyl regulates AAA formation and showed that circCdyl promotes vascular inflammation and M1 polarization by inhibiting interferon regulatory factor 4 (IRF4) entry into the nucleus, significantly inducing AAA formation. In addition, circCdyl was shown to act as a let-7c sponge, promoting C/EBP-δ expression in macrophages to induce M1 polarization. Our results indicate an important role for circCdyl-mediated macrophage polarization in AAA formation and provide a potent therapeutic target for AAA treatment.

Achalasia and acromegaly: Co-incidence of these diseases or a new syndrome?

BACKGROUND: Acromegaly is a disorder associated with hypersecretion of growth hormone, most usually caused by a pituitary adenoma. Dysmotility of the gastrointestinal tract has been reported in acromegalic patients. Achalasia is a disorder characterized by aperistalsis of the oesophagus with incomplete lower oesophageal sphincter relaxation and whose aetiology remains unknown. Mutations in some genes have previously been associated with the development of acromegaly or achalasia. The study aims were to analyse mutations in selected genes in a woman having both of these diseases, to identify their aetiological factors, and to suggest explanations for the co-incidence of acromegaly and achalasia. METHODS AND RESULTS: A female patient with acromegaly, achalasia, and a multinodular thyroid gland with hyperplastic colloid nodules underwent successful treatment of achalasia via laparoscopic Heller myotomy, a thyroidectomy was performed, and the pituitary macroadenoma was surgically excised via transnasal endoscopic extirpation. Germline DNA from the leukocytes was analysed by sequencing methods for a panel of genes. No pathogenic mutation in AAAS, AIP, MEN1, CDKN1B, PRKAR1A, SDHB, GPR101, and GNAS genes was found in germline DNA. The somatic mutation c.601C>T/p.R201C in the GNAS gene was identified in DNA extracted from a tissue sample of the pituitary macroadenoma. CONCLUSIONS: We here describe the first case report to our knowledge of a patient with both acromegaly and achalasia. Association of acromegaly and soft muscle tissue hypertrophy may contribute to achalasia's development. If one of these diagnoses is determined, the other also should be considered along with increased risk of oesophageal and colorectal malignancy.

Protein signature of human skin fibroblasts allows the study of the molecular etiology of rare neurological diseases.

BACKGROUND: The elucidation of pathomechanisms leading to the manifestation of rare (genetically caused) neurological diseases including neuromuscular diseases (NMD) represents an important step toward the understanding of the genesis of the respective disease and might help to define starting points for (new) therapeutic intervention concepts. However, these "discovery studies" are often limited by the availability of human biomaterial. Moreover, given that results of next-generation-sequencing approaches frequently result in the identification of ambiguous variants, testing of their pathogenicity is crucial but also depending on patient-derived material. METHODS: Human skin fibroblasts were used to generate a spectral library using pH8-fractionation of followed by nano LC-MS/MS. Afterwards, Allgrove-patient derived fibroblasts were subjected to a data independent acquisition approach. In addition, proteomic signature of an enriched nuclear protein fraction was studied. Proteomic findings were confirmed by immunofluorescence in a muscle biopsy derived from the same patient and cellular lipid homeostasis in the cause of Allgrove syndrome was analysed by fluorescence (BODIPY-staining) and coherent anti-Stokes Raman scattering (CARS) microscopy. RESULTS: To systematically address the question if human skin fibroblasts might serve as valuable biomaterial for (molecular) studies of NMD, we generated a protein library cataloguing 8280 proteins including a variety of such linked to genetic forms of motoneuron diseases, congenital myasthenic syndromes, neuropathies and muscle disorders. In silico-based pathway analyses revealed expression of a diversity of proteins involved in muscle contraction and such decisive for neuronal function and maintenance suggesting the suitability of human skin fibroblasts to study the etiology of NMD. Based on these findings, next we aimed to further demonstrate the suitability of this in vitro model to study NMD by a use case: the proteomic signature of fibroblasts derived from an Allgrove-patient was studied. Dysregulation of paradigmatic proteins could be confirmed in muscle biopsy of the patient and protein-functions could be linked to neurological symptoms known for this disease. Moreover, proteomic investigation of nuclear protein composition allowed the identification of protein-dysregulations according with structural perturbations observed in the muscle biopsy. BODIPY-staining on fibroblasts and CARS microscopy on muscle biopsy suggest altered lipid storage as part of the underlying disease etiology. CONCLUSIONS: Our combined data reveal that human fibroblasts may serve as an in vitro system to study the molecular etiology of rare neurological diseases exemplified on Allgrove syndrome in an unbiased fashion.