Intracellular Enzyme Research Articles

Serum and Cerebrospinal Fluid Lactate Dehydrogenase in Children with Febrile Convulsions.

Tissue damage caused by febrile convulsion has not still been proved or refuted completely. Given the fact that lactate dehydrogenase as an intracellular enzyme can be increased due to tissue damage, we decided to evaluate serum and cerebrospinal fluid lactate dehydrogenase in children with febrile convulsion. This is a cross-sectional study on 166 children aged 6-24 month, in three groups of simple febrile convulsion (n=56), complex febrile convulsion (n=27) with 3 different subgroups (recurrence in 24 hours, duration >15 minutes, and with focal components), and control (n=83). Patients' serum and cerebrospinal fluid specimens were collected after meeting the inclusion criteria. Demographic information was documented and patients' serum and cerebrospinal fluid lactate dehydrogenase and glucose were measured. Data were analyzed using SPSS software. The mean serum lactate dehydrogenase in simple febrile convulsion, complex febrile convulsion, and controls were 501.57± 143.70, 553.07±160.22, and 505.87±98.73 U/L, respectively. The mean cerebrospinal fluid lactate dehydrogenase in simple, complex febrile convulsion, and control groups were 22.58±11.92, 29.48±18.18, and 21.56±17.32 U/L, respectively. Only cerebrospinal fluid lactate dehydrogenase difference between complex febrile convulsion and control group (p=0.039) (In the duration >15 minutes subgroup and controls, p=0.028) was statistically significant. There was a significant difference between sex and serum lactate dehydrogenase in thesame subgroup of complex group (p=0.012). Complex febrile convulsion may lead to increase of lactate dehydrogenase in cns of CNS cellular damage.

Antioxidant and Antiapoptotic Properties of n-Butanol Fraction the Acanthopanax senticosus Extracts in H2O2-RAW264.7 Cells and CCl4-Induced Liver Injury in Mice.

The Acanthopanax senticosus has been shown to have a wide range of pharmacological activities, which are associated with health benefits, such as antioxidant, anti-inflammatory, and antiapoptotic properties. A previous study has shown that the n-butanol fraction of A. senticosus extract had the strongest antioxidant effect in vitro. This study aimed to investigate the effects that the n-butanol fraction of A. senticosus extract could relieve oxidative stress damage through antioxidant and antiapoptotic in the H2O2-stimulated RAW264.7 macrophages and the CCl4-induced liver injury. The result showed that the n-butanol fraction extract could relieve damage by increasing the intracellular antioxidant enzymes (SOD) level, decreasing intracellular ROS and MDA levels, and regulating antioxidant and antiapoptotic-related gene expression levels. The morphological observation of HE, TUNE, and immunohistochemistry staining of liver tissue verified that the n-butanol fraction extract is though anti-oxidative and antiapoptotic to alleviate cellular oxidative damage. The RT-PCR assay showed that the Keap1-Nrf2-ARE and the Bax/Bcl-2 signaling pathway were related to the molecular mechanism of action. The experimental results show that Acanthopanax senticosus extract has a good effect in treating liver injury and enhancing the antioxidant capacity of the body.

Synthesis of NMN by cascade catalysis of intracellular multiple enzymes.

β-Nicotinamide mononucleotide is a biologically active nucleotide compound, and its excellent anti-aging activity is widely used in medicine and has multiple functions, making NMN have broad application prospects in the fields of nutrition, health food, and even medicine. Herein, based on the supply of the co-substrate PRPP, we designed and constructed three in vivo NMN synthesis pathways using glucose, xylose, and arabinose as raw materials and Escherichia coli as the host. The best in vivo pathway through whole-cell catalysis was identified. Then, we optimized the cell culture and catalytic conditions of the optimal path to determine the optimal conditions and ultimately obtained an NMN titer of 1.8 mM.

RNA Dysmetabolism and Repeat-Associated Non-AUG Translation in Frontotemporal Lobar Degeneration/Amyotrophic Lateral Sclerosis due to C9orf72 Hexanucleotide Repeat Expansion.

Neuropathological features of frontotemporal dementia and amyotrophic lateral sclerosis (ALS) due to C9orf72 GGGGCC hexanucleotide repeat expansion include early dipeptide repeats, repeat RNA foci, and subsequent TDP-43 pathologies. Since the discovery of the repeat expansion, extensive studies have elucidated the disease mechanism of how the repeat causes neurodegeneration. In this review, we summarize our current understanding of abnormal repeat RNA metabolism and repeat-associated non-AUG translation in C9orf72 frontotemporal lobar degeneration/ALS. For repeat RNA metabolism, we specifically focus on the role of hnRNPA3, the repeat RNA-binding protein, and the EXOSC10/RNA exosome complex, an intracellular RNA-degrading enzyme. In addition, the mechanism of repeat-associated non-AUG translation inhibition via TMPyP4, a repeat RNA-binding compound, is discussed.

Creation and Characterization of Mycolicibacterium Smegmatis mc2155 with Deletions in Genes Encoding Sterol Oxidation Enzymes

The fast-growing saprotrophic strain Mycolicibacterium smegmatis mc2155 is capable of utilizing plant and animal sterols and can be used for creation of genetically engineered strains producing biologically active steroids. Oxidation of the 3β-hydroxyl group and Δ5(6)→Δ4(5) double bond isomerization followed by formation of stenones from sterols are considered as the initial stage of steroid catabolism in some actinobacteria. The study of the mechanism of steroid nucleus 3β-hydroxyl group oxidation is relevant for the creation of a method of the microbiological production of valuable 3β-hydroxy-5-en-steroids. A mutant strain of M. smegmatis with deletions in three genes (MSMEG_1604, MSMEG_5228 and MSMEG_5233) encoding known enzymes exhibiting 3β-hydroxysteroid dehydrogenase activity was constructed by homologous recombination coupled with double selection. The resulting mutant retained macromorphological properties and the ability to convert cholesterol. 3-Keto-4-en-steroids were found among the sterol catabolism intermediates. Experimentally obtained data indicate the presence of a previously undetected intracellular enzyme that performs the function of 3β-hydroxysteroid dehydrogenase/Δ5(6)→Δ4(5) isomerase.

Apremilast in Patients Difficult to Treat: A Multicentric Real‐Life Long‐Term Experience

Psoriasis is an inflammatory disease nowadays considered not only as a cutaneous but also as a systemic disease. Systemic therapy plays a crucial role in the management of psoriasis. Apremilast is an inhibitor of phosphodiesterase‐4 (PDE4), indicated in the treatment of moderate‐to‐severe psoriasis. Here, we report a multicentric case series of patients treated with apremilast with resolution of skin manifestations and maintenance of clinical response for a minimum of 2 years. By inhibiting PDE4, apremilast acts as a ubiquitous intracellular enzyme, whose active form degrades adenosine cyclic intracellular monophosphate (cAMP) into AMP. The increase in cAMP determines a decrease in proinflammatory cytokines such as TNF‐a, IL‐17, IL‐23, and upregulation of IL‐10 with an anti‐inflammatory action. Considering the growing incidence of comorbidities in the world population and in particular the strict correlation in patients with psoriasis, it is important to identify therapeutic options able to avoid a negative impact on patients with both conditions. The aim of this work is to highlight the utility of this molecule in the long‐term management of these patients. Moreover, these case series further underline the high safety profile and manageability of this small molecule.

WSL214 negatively regulates ROS accumulation and pathogen defense response in rice.

WSL214 plays an important role in promoting cellular ROS homeostasis by enhancing catalase activity and reducing photosynthetic ROS production. ROS are the important regulator of cellular homeostasis, and balancing ROS production and clearance contributes to cellular activity. Although many genes associated with ROS have been cloned, the mechanism of this balance is not fully understood. In this study, we obtained the rice mutant wsl214 that arose from a natural mutation. Compared to WT, wsl214 exhibited white-striped leaves, defective chloroplast development, reduced net photosynthetic rate, and overexcitation of photosynthetically active reaction centers. In addition, the ROS accumulation level was significantly elevated, and the ROS scavenging enzyme activity was significantly decreased in wsl214 leaf tissue. As a result of elevated ROS levels, wsl214 leaf cells underwent DNA damage and programmed cell death. However, wsl214 defense response to exogenous pathogens was also enhanced by high ROS levels. Based on the mapping cloning, we discovered that WSL214 had a single base mutation (C to T) in the third exon, resulting in decreased expression of wsl214. The WSL214 encodes an HD domain phosphohydrolase and is widely expressed in various tissues of rice, especially at the highest level in leaf tissue. Further research showed that WSL214 promoted the homeostasis of rice leaf cellular ROS in two ways. First, WSL214 increased the expression of the catalase gene OsCATC, making the intracellular ROS scavenging enzyme more active.Second, WSL214 promoted chloroplast development, kept photosynthesis working properly, and reduced ROS produced by photosynthesis.In conclusion, our report emphasizes that WSL214 is a key part of balancing ROS levels in cells.

STATE OF THE PHAGOCYTIC LINK OF IMMUNITY IN CHOLANGITIS

Summary. Aim. To study the condition of the phagocytic part of immunity in cholangitis. One of the main ways of elimination of bacterial contamination of the body is the work of the immune system, namely phagocytosis. This especially applies to acute and chronic cholangitis.
 Materials and methods. Depending on the magnitude of pathological changes in the biliary system, we have identified three groups of patients in whom cholangitis occurs in conditions that progress and worsen. Group I – cholangitis in conditions of choledocholithiasis, group II – cholangitis in conditions of iatrogenic strictures of the main bile ducts, and group III – cholangitis in conditions of already performed reconstruction of the duct system. The functional activity of phagocytic cells was studied.
 Results and their discussion. It was established that the functional capabilities of neutrophils are depleted in proportion to the severity of cholangitis and its duration. In case of chronic undulating cholangitis, there is an insufficiency of the phagocytic link, phagocytes work on the verge of exhausting their reserve capabilities. Adhesion forces and the potential of intracellular lysosomal enzymes are especially affected.
 Conclusions. The functional capacity of phagocytic cells in cholangitis is characterized by a deficiency, the degree of virulence is to lie down in the severity of inflammation and thrive. The greatest manifestations of changes were revealed in acid-independent mechanisms of phagocytosis, which were characterized by impaired adhesion and the release of intracellular lysosomal enzymes.

Study of the Working Parameters of the Enzyme Protepsin and Its Effect on The Microstructure of Second-Grade Beef

"Protepsin" is an enzyme preparation of animal nature containing a complex of acid proteinases, intended for use in the meat industry for processing meat raw materials. The enzyme composition of the preparation is balanced according to the degree of exposure to various proteins of meat and meat systems used in the technology of obtaining meat products. "Protepsin" works in the meat system similarly to intracellular enzymes (cathepsins). It is their synergist and has additional qualities that allow it to act in a wider range of technological parameters, as well as to influence those protein systems on which intracellular enzymes do not act or have an effect to an insignificant extent [1]. The practical significance of the work lies in the fact that semismoked sausage products are in particular demand among the population. Cost reduction while maintaining their quality in a competitive environment is a time requirement. Therefore, the use of the enzyme is one of the promising directions in the field of meat processing [1]. The aim of the work is to study the working parameters of the enzyme "Protepsin" and its effect on the microstructure of second-grade beef. Histological changes characterize the positive effect of the enzyme on the development of meat quality indicators. In this regard, the processing of meat raw materials with "Protepsin" leads to a significant reduction in the duration of maturation of meat, which makes it possible to achieve the necessary changes in a shorter time compared with the natural way of autolysis.

Characterization of cell death in harmful Karenia mikimotoi under algicidal activity of Marinobacter sp. O-7

Use of algicidal bacteria, which has the advantage of both safety and efficiency, has gradually become a significant strategy to handle the environmental problem of Harmful Algal Blooms (HABs). A strain of bacterium with excellent algicidal activity of 95.6% and identified as Marinobacter sp. (O-7) was isolated from seawater. The results demonstrated that strain O-7 can secrete active substances, which can pose a fatal threat to harmful Karenia mikimotoi. The morphology of K. mikimotoi cells changed over time. The physiological responses of algal cells after exposure to strain O-7 algicidal compounds were as follows: photosynthetic pigments (chlorophyll a and carotenoid) levels of the algal cells decreased significantly as did the photosynthetic efficiency and performance index, while reactive oxygen species levels substantially increased under the stress of 3% dose of O-7 for 4 h of treatment. Concurrently, the intracellular antioxidant enzymes system (superoxide dismutase, catalase, glutathione peroxidase) and non-enzymes system (glutathione, oxidized glutathione, ascorbic acid and dehydroascorbic acid) showed up-regulation. The content of malondialdehyde amplified with increase in concentration, indicating lipid peroxidation of algal cells. Collectively, our results demonstrated that the secretion of O-7 could act as an effective algicide for the control of harmful algae and offered clues to understanding the death mechanism of indirect processing of bacterial secretion against microalgae.

Aerobic Degradation Characteristics and Mechanism of Decabromodiphenyl Ether (BDE-209) Using Complex Bacteria Communities.

Complex bacteria communities that comprised Brevibacillus sp. (M1) and Achromobacter sp. (M2) with effective abilities of degrading decabromodiphenyl ether (BDE-209) were investigated for their degradation characteristics and mechanisms under aerobic conditions. The experimental results indicated that 88.4% of 10 mg L-1 BDE-209 could be degraded after incubation for 120 h under the optimum conditions of pH 7.0, 30 °C and 15% of the inoculation volume, and the addition ratio of two bacterial suspensions was 1:1. Based on the identification of BDE-209 degradation products via liquid chromatography-mass spectrometry (LC-MS) analysis, the biodegradation pathway of BDE-209 was proposed. The debromination, hydroxylation, deprotonation, breakage of ether bonds and ring-opening processes were included in the degradation process. Furthermore, intracellular enzymes had the greatest contribution to BDE-209 biodegradation, and the inhibition of piperyl butoxide (PB) for BDE-209 degradation revealed that the cytochrome P450 (CYP) enzyme was likely the key enzyme during BDE-209 degradation by bacteria M (1+2). Our study provided alternative ideas for the microbial degradation of BDE-209 by aerobic complex bacteria communities in a water system.

Effect of the antagonistic yeast Hannaella sinensis on the degradation of Patulin

Patulin (PAT) is a toxic fungal metabolite that is widely distributed in food and poses a serious health risk to humans and animals. Hannaella sinensis has been found to be effective in controlling post-harvest diseases of apples. However, it is not known whether H. sinensis can degrade PAT. The aim of this study was to investigate the degradation efficacy of PAT by H. sinensis and the possible mechanisms involved. The experimental results showed that H. sinensis degraded 100 % of PAT in NYDB medium within 30 h. The living cells of H. sinensis did not absorb PAT, and the cell wall and extracellular metabolites of H. sinensis were not involved in the degradation of PAT. The addition of cycloheximide to the culture medium inhibited the degradation of PAT by H. sinensis. Our results indicated that H. sinensis could degrade PAT and that the intracellular enzymes of H. sinensis played a crucial role in the degradation process. This result provides a new solution for the degradation of PAT.

Inclusion of catalase in semen extender modulates the post thaw semen quality and oxidative stress profiles in mithun spermatozoa

Present study was designed to assess the effect of catalase on post-thaw semen quality parameters (SQPs), sperm kinematics, antioxidant and oxidative stress profiles and sperm cholesterol efflux in mithun (Bos frontalis). A total of 25 ejaculates were selected based on biophysical parameters and each sample was split into four equal aliquots and diluted (to get final concentration of 60 million spermatozoa per mL) with the TCG extender containing either 0 U/mL, 50 U/mL, 100 U/mL, and 150 U/mL catalase in the Group I, II, III and IV, respectively. Frozen-thawed samples were analysed for motility parameters (progressive forward and in bovine cervical mucus [BCMPT]), kinetic and velocity parameters by computer-assisted sperm analyser (CASA), viability, sperm morphological and nuclear abnormalities, acrosomal integrity, plasma membrane and nuclear integrities, sperm intra-cellular enzymatic leakage and seminal plasma biochemical (sperm cholesterol and oxidative stress markers) profiles. Study revealed an enhancement in viability, acrosomal integrity, plasma membrane integrity, motility (progressive and in cervical mucus), sperm cholesterol content and reduction in sperm morphological and nuclear abnormalities, leakage of intracellular enzymes in Group III. Moreover, intactness of acrosome and biochemical membranes were protected significantly in addition to significant improvement in kinetic and velocity profiles in extender containing 100 U/ml catalase. Correlation analysis revealed that sperm kinetic parameters, SQPs and antioxidant parameters had significant positive correlation with each other whereas these profiles were negatively correlated with sperm morphological abnormalities, enzymatic leakage and lipid peroxidation in catalase treated sperm. The results clearly indicated that inclusion of 100 U/ml catalase holds a clear advantage over control or 50 U/ml or 150 U/ml catalase in cryopreservation of mithun semen. It can be concluded from the present study that catalase supplementation in semen extender can be effectively utilized to reduce the oxidative stress and to improve the post-thaw semen quality in mithun.

Markers for internal neoplasia in the horse

The diagnosis of internal neoplasia in horses is challenging. Increased production of hormones physiologic for adult animals (e.g., adrenocorticotropin, norepinephrine, and erythropoietin) or typical for the foetal phase (alpha-fetoprotein, anti-Müllerian hormone, and parathyroid-hormone-related protein) might aid in tumour diagnostics. Thymidine kinase-1 and alkaline phosphatase are examples of intracellular enzymes, whose activity in the blood may increase in some neoplasia cases. Furthermore, inappropriate production of abnormal monoclonal or autologous antibodies can accompany lymphoma and multiple myeloma. Many of those tumour markers lead to clinical or laboratory changes, called paraneoplastic syndromes, such as hypercalcaemia and erythrocytosis. The interpretation of the results of the tumour marker measurements in horses is complicated due to many factors affecting the markers' concentration or activity (e.g., young age, pregnancy, and inflammation) and other diseases triggering the same changes. Moreover, the presence of paraneoplastic syndromes is inconsistent, which leads to low sensitivity of those substances as tumour markers. In conclusion, screening for neoplasia in horses is not recommended. The measurement of tumour markers should be performed only in risk groups with suspicious clinical or laboratory findings, and the results should be interpreted with caution. It is advisable to add inflammatory markers to the tumour profile or repeat the measurements.

Phosphodiesterase 5 Inhibitor Potentiates Epigallocatechin 3-O-Gallate-Induced Apoptotic Cell Death via Activation of the cGMP Signaling Pathway in Caco-2 Cells.

Epigallocatechin 3-O-gallate (EGCG) is a predominant component in green tea with various health benefits. The 67 kDa laminin receptor (67LR) is a nonintegrin cell surface receptor that is overexpressed in various types of cancer; 67LR was identified a cell surface EGCG target that plays a pivotal role in tumor growth, metastasis, and resistance to chemotherapy. However, the plasma concentration of EGCG is limited, and its molecular mechanisms remain unelucidated in colon cancer. In this study, we found that the phosphodiesterase 5 (PDE5) inhibitor, vardenafil (VDN), potentiates EGCG-induced apoptotic cell death in colon cancer cells. The combination of EGCG and VDN induced apoptosis via activation of the endothelial nitric oxide synthase/cyclic guanosine monophosphate/protein kinase Cδ signaling pathway. In conclusion, the PDE5 inhibitor, VDN, may reduce the intracellular PDE5 enzyme activity that potentiates EGCG-induced apoptotic cell death in Caco-2 cells. These results suggest that PDE5 inhibitors can be used to elevate cGMP levels to induce 67LR-mediated, cancer-specific cell death. Therefore, EGCG may be employed as a therapeutic candidate for colon cancer.

A New Glutathione-Cleavable Theranostic for Photodynamic Therapy Based on Bacteriochlorin e and Styrylnaphthalimide Derivatives

Herein, we report a new conjugate BChl-S-S-NI based on the second-generation photosensitizer bacteriochlorin e6 (BChl) and a 4-styrylnaphthalimide fluorophore (NI), which is cleaved into individual functional fragments in the intracellular medium. The chromophores in the conjugate were cross-linked by click chemistry via a bis(azidoethyl)disulfide bridge which is reductively cleaved by the intracellular enzyme glutathione (GSH). A photophysical investigation of the conjugate in solution by using optical spectroscopy revealed that the energy transfer process is realized with high efficiency in the conjugated system, leading to the quenching of the emission of the fluorophore fragment. It was shown that the conjugate is cleaved by GSH in solution, which eliminates the possibility of energy transfer and restores the fluorescence of 4-styrylnaphthalimide. The photoinduced activity of the conjugate and its imaging properties were investigated on the mouse soft tissue sarcoma cell line S37. Phototoxicity studies in vitro show that the BChl-S-S-NI conjugate has insignificant dark cytotoxicity in the concentration range from 15 to 20,000 nM. At the same time, upon photoexcitation, it exhibits high photoinduced activity.

Tetrahydroxy Stilbene Glucoside Alleviates Ischemic Stroke by Regulating Conformation-Dependent Intracellular Distribution of PKM2 for M2 Macrophage Polarization.

Tetrahydroxy stilbene glucoside (TSG) is a bioactive ingredient with powerful anti-inflammatory and neuroprotective properties. However, the detailed mechanisms concerning the neuroprotective effect of TSG are not fully understood. This study aims to address the molecular mechanism involved in the protective effects of TSG on murine ischemic stroke. We found that TSG meliorated the phenotypes of ischemic stroke in vivo, which was correlated with the increased percentage of infiltrated M2 macrophages in brain after stroke. Mechanistically, TSG regulated macrophage polarization by significantly downregulating the transcriptional levels of M1 marker genes (iNOS and IL-1β) but upregulating that of the M2 marker genes (arg-1 and IL-4) following lipopolysaccharide/interferon-γ stimulation. Consistently, TSG reversed the metabolic profiling of M1 macrophage toward the M2 status at intracellular energy levels. Surprisingly, the knockdown of an established metabolic enzyme pyruvate kinase M2 (PKM2) that is important for M1 switch in macrophages abolished the promotive effect of TSG on the M2 polarization. Further investigation revealed that TSG markedly downregulated the intracellular ratio of dimer/monomer to the tetramer of PKM2 without affecting its total protein expression, leading to a suppressed nuclear translocation of functioning PKM2 in macrophages for M1 differentiation. Taken together, we identified a novel mechanism for macrophage M2 polarization regulation by a small-molecule chemical that controls the quality (conformation) rather than the quantity (expression) of an intracellular M1-promoting metabolic enzyme, which offers a better understanding of the mechanisms of macrophage plasticity and has serious implication in translational strategies for the treatment of macrophage-mediated neurological diseases with natural bioactive products.

Inflammation-related citrullination of matrisome proteins in human cancer.

Protein arginine deiminases (PADs) are intracellular enzymes that may, especially in pathological conditions, also citrullinate extracellular substrates, including matrisome proteins such as structural proteins in extracellular matrix (ECM). PADs are abundantly expressed in human cancer cells. Citrullination of matrisome proteins has been reported in colon cancer but the phenomenon has never been systematically studied. To gain a broader view of citrullination of matrisome proteins in cancer, we analyzed cancer proteomics data sets in 3 public databases for citrullinated matrisome proteins. In addition, we used three-dimensional cell cocultures of fibroblasts and cancer cells and analyzed citrullination of ECM. Our new analysis indicate that citrullination of ECM occurs in human cancer, and there is a significant variation between tumors. Most frequently citrullinated proteins included fibrinogen and fibronectin, which are typically citrullinated in rheumatoid inflammation. We also detected correlation between immune cell marker proteins, matrix metalloproteinases and ECM citrullination, which suggests that in cancer, citrullination of matrisome proteins is predominantly an inflammation-related phenomenon. This was further supported by our analysis of three-dimensional spheroid co-cultures of nine human cancer cell lines and fibroblasts by mass spectrometry, which gave no evidence that cancer cells or fibroblasts could citrullinate matrisome proteins in tumor stroma. It also appears that in the spheroid cultures, matrisome proteins are protected from citrullination.

Amylolytic activity of <i>Bacillus subtilis</i> isolates obtained from wildlife microbiota

At present, the study of the enzymatic potential of Bacillus subtilis bacteria is in demand due to the fact that probiotic and enzymatic preparations are created on their basis for their use in animal husbandry. The results of the study of amylolytic activity of 10 isolates of B. subtilis bacteria isolated from wildlife microbiota for amylopectin are presented. Determination of bacterial enzyme activity was performed by the Schomodi-Nelson method. The supernatants of the culture liquid of the 1st, 2nd, 3rd, and 7th day cultures were taken as enzyme preparation. Isolates with high qualitative indices were examined quantitatively using a spectrophotometer. The two isolates with the best performance were selected. Protein concentration was determined by the Bradford method. In addition to studying supernatants of isolates that showed extracellular enzymatic activity, studies were performed on intracellular enzymes. Cell destruction was performed using an ultrasonic disintegrator, in which the dissolved sediment was contained in a glass vial in a flask with ice. The best qualitative indicators were found in B. subtilis 2SP and B. subtilis 5SP isolates, which had high amylopectin activity from day one, so further quantitative studies were continued with them. Protein concentrations in the above isolates progressed by the days of seeding. The specific activity also increased accordingly and on day 7, the B. subtilis 2SP isolate was 2.75 units/mg, B. subtilis 5SP 2.67 units/mg. The study of the activity of intracellular enzymes in these isolates confirmed similar qualitative indicators. Two B. subtilis isolates were selected as promising for the development of enzyme preparations isolated from the microbiota of wild animals of the Far North.

Platycodin D restores the intestinal mechanicalbarrier by reducing endoplasmic reticulum stress-mediated apoptosis

• Exploring the regulatory effect of PD on CP-induced intestinal injury. • Demonstrating that PD exerts a certain protective effect on intestinal damage caused by CP. • Revealing that PD for alleviating CP-induced intestinal damage might be related to PERK-eIF2ɑ-ATF4 signaling pathway. The current study aimed to reveal the inhibitory effect of Platycodin D (PD) on intestinal cell apoptosis mediated by ER stress. Treatment with PD alleviated pathological changes of the intestine and reduced secretion of intracellular enzyme Diamine oxidase (DAO) in the upper villi of the small intestinal mucosa in Cisplatin (CP) intestinal injury model and increased expression of key protein Occludin and ZO-1 of the intestinal mechanical barrier. PD inhibited ER stress in a dose-dependent in CP-caused IEC-6 cells and the level of phosphorylation of Protein kinase R-like ERK kinase (PERK) and Eukaryotic initiation factor-2 alpha (eIF2α) reduced. Besides, the ER stress inhibitor 4-PBA was used to confirm that PD alleviated the ER stress in vitro . In conclusion, PD exerted an intestinal protective effect through the inhibition of ER stress in CP-treated mice and IEC-6 cells. These data indicated that PD was a promising natural medicine for intestinal protection.