Lactate Dehydrogenase Enzyme Activity Research Articles

Hazelnut Skin in Ewes' Diet: Effects on Colostrum Immunoglobulin G and Passive Transfer of Immunity to the Lambs.

Passive immunity transfer has a pivotal role in newborn lambs, where the colostrum represents the primary source of immunoglobulins. This study hypothesized that the high content in polyphenolic compounds, mono- and poly-unsaturated fatty acids, and vitamin E of hazelnut skin affects blood and colostrum immunoglobulin G (IgG) concentration and related gamma-glutamyl-transferase (GGT) and lactate dehydrogenase (LDH) levels in sheep and their lambs. In the last 45 days of pregnancy, ewes were divided into a control (CTR) and a hazelnut skin supplemented group (HZN). Blood and colostrum were collected from ewes and lambs before the first suckling, at 24 and 48 h after birth, then IgG concentration, GGT and LDH activity levels were measured. IgG concentration in the colostrum and in lamb's serum were significantly greater in HZN than CTR. No significant difference was detected for ewe's blood. A significant positive correlation was found between IgG and GGT in lambs' serum and colostrum, between IgG and LDH, as well as between GGT and LDH in lambs' serum and colostrum. Our results suggest that hazelnut skin supplementation influences IgG colostrum concentration, with improved immune passive transfer to the suckling lambs. The transfer of maternal derived immune factors is confirmed by the GGT and LDH enzyme activity levels.

L-carnitine modulates free mitochondrial DNA DAMPs and platelet storage lesions during storage of platelet concentrates.

Platelet storage lesions may occur in Platelet concentrates (PCs) storage time, reducing PCs' quality. Mitochondrial damage causes mitochondrial DNA (mtDNA) to be released into the extracellular space. In this study, we evaluated the effect of L-carnitine (LC) as an antioxidant on free mtDNA DAMPs release in PCs during storage. Ten PCs prepared by the PRP method were studied. The copy numbers of free mtDNA, total reactive oxygen species (ROS), lactate dehydrogenase (LDH) enzyme activity, pH, and platelet counts were measured on days 0, 3, 5, and 7 of PCs storage in LC-treated and untreated platelets. LDH activity was significantly lower than the control group during 7 days of PCs storage (p = 0.041). Also, ROS production decreased in LC-treated PCs compared to the control group during storage (p = 0.026), and the difference mean of ROS between the two groups was significant on day 3, 5, and 7 (Pday3 = 0.02, Pday5 = 0.0001, Pday7 = 0.031). Moreover, LC decreased the copy numbers of free mtDNA during 7 days of storage (p = 0.021), and the difference mean of the copy numbers of free mtDNA in LC-treated PCs compared to the control group was significant on day 5 and 7 (Pday5 = 0.041، Pday7 = 0.022). It seems that LC can maintain the metabolism and antioxidant capacity of PCs and thus can reduce mitochondrial damage and mtDNA release; consequently, it can decrease DAMPs in PCs. Therefore, it may be possible to use this substance as a platelet additive solution in the future.

Alantolactone induces concurrent apoptosis and GSDME-dependent pyroptosis of anaplastic thyroid cancer through ROS mitochondria-dependent caspase pathway

BackgroundAnaplastic thyroid cancer (ATC) is one of the fatal cancers and has not effective treatments. Alantolactone (ATL), a terpenoid extracted from traditional Chinese medicinal herb Inula helenium L., confers significant anti-inflammatory, antibacterial and antitumor activity. However, the activity and mechanisms of ATL in ATC remain unclear. PurposeTo investigate the potential anti-ATC effects in vitro and in vivo and the mechanisms involved. MethodsThe anti-proliferative activity of Alantolactone (ATL) against ATC cells was analyzed through CCK-8 and colony formation assays. Flow cytometry assay was performed to assess the cell cycle, cell apoptosis, ROS, and mitochondrial membrane potential (ΔΨm), whereas the cellular localization of cytochrome c and calreticulin were determined using cellular immunofluorescence assays. The lactate dehydrogenase (LDH) enzyme activity in the cell culture medium was measured using a commercial LDH kit, whereas ELISA was conducted to assess the secretory function of IL-1β. Western blot assays were conducted to determine the expression or regulation of proteins associated with apoptosis and pyroptosis. Subcutaneous tumor model of nude mice was established to evaluate the anticancer activity of ATL in vivo. The expression of Ki67, cyclin B1, cleaved-PARP, cleaved-caspase 3, and IL-1β in the animal tumor tissues was profiled using immunohistochemistry analyses. ResultsOur data showed that ATL significantly inhibited the proliferation and colony formation activity of ATC cells. ATL induced ATC cell cycle arrest at G2/M phase, and downregulated the expression of cyclin B1 and CDC2. Furthermore, ATL induced concurrent apoptosis and pyroptosis in the ATC cells, and the cleavage of PARP and GSDME. It also significantly increased the release of LDH and IL-1β. Mechanically, ATL-mediated increase in ROS suppressed the Bcl-2/Bax ratio, downregulated the mitochondrial membrane potential and increased the release of cytochrome c, leading to caspase 9 and caspase 3 cleavage. We also found that ATL induced the translocation of an immunogenic cell death marker (calreticulin) to the cell membrane. In addition, it inhibited the growth of the ATC subcutaneous xenograft model, and activated proteins associated with apoptosis and pyroptosis, with a high safety profile. ConclusionTaken together, these results firstly demonstrated that ATL exerted an anti-ATC activity by inducing concurrent apoptosis and GSDME-dependent pyroptosis through ROS-mediated mitochondria-dependent caspase activation. Meanwhile, these cell deaths exhibited obvious characteristics of immunogenic cell death, which may synergistically increase the potential of cancer immunotherapy in ATC. Further studies are needed to explore deeper mechanisms for the anti- ATC activity of ATL.

TGF-β3 induces lactate production in Sertoli cell through inhibiting the Notch pathway.

In the testis, spermatocytes and spermatids rely on lactate produced by Sertoli cells (SCs) as energy source. Transforming growth factor-beta 3 (TGF-β3) is one of the generally accepted paracrine regulatory factors of SC-created blood-testis barrier (BTB), yet its role in SC glycolysis and lactate production still remains unclear. To investigate the effect of TGF-β3 on glycolysis and lactate production in SCs and determine the role of lethal giant larvae 2 (Lgl2) and Notch signaling activity during this process. Primary cultured rat SCs and TM4 cells were treated with different concentrations of TGF-β3. In some experiments, cells were transfected with siRNA specifically targeting Lgl2 and then treated with TGF-β3 or N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester. Lactate concentration, glucose and glutamine (Gln) consumption in the culture medium, activity of phosphofructokinase (PFK), lactate dehydrogenase (LDH), and glutaminase (Gls), ATP level, oxygen consumption, extracellular acidification, and mitochondrial respiration complex activity were detected using commercial kits. The protein level of Lgl2, LDH, monocarboxylate transporter 4 (MCT4), and activity of Akt, ERK, p38 MAPK, and Notch pathway were detected by Western blot. The stage-specific expression of Jagged1 was examined by immunohistochemistry (IHC) and qPCR after laser capture microdissection. Spermatogenesis in rat testis injected with recombinant Jagged1 (re-Jagged1) was observed by HE staining, and lactate concentration in testis lysate was measured at a different day point after re-Jagged1 treatment. Significant enhancement of lactate concentration was detected in a culture medium of both primary SCs and TM4 cells treated with TGF-β3 at 3 or 5ng/ml. Besides, other parameters of glycolysis, that is, glucose and Gln consumption, enzyme activity of PFK, LDH, and Gls displayed different levels of increment in primary SCs and TM4 cells after TGF-β3 treatment. Mitochondria respiration of SCs was shown to decrease in response to TGF-β3. Lgl2, MCT4, activity of ERK, and p38 MAPK were up-regulated, whereas Akt and Notch pathway activity were inhibited by TGF-β3. Silencing of Lgl2 in SCs affected lactate production and attenuated the previous effects of TGF-β3 on SC glycolysis except for Gln consumption, Gls activity, and activity of Akt, ERK, and p38. N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT) treatment in SCs antagonized glycolysis suppression caused by Lgl2-silencing. In vivo analysis revealed a stage-specific expression of Jagged1 in contrary with TGF-β3. Activating Notch signaling by re-Jagged1 resulted in restorable hypospermatogenesis and lowered lactate level in rat testis. TGF-β3 induces lactate production in SC through up-regulating Lgl2, which weakened the Notch signaling activity and intensified glycolysis in SCs. Thus, besides the known function of TGF-β3 as the BTB regulator, TGF-β3-Lgl2-Notch may be considered an important pathway controlling SC glycolysis and spermatogenesis.

Fluted pumpkin seeds protect the spermatogenesis score index and testicular histology of caffeine treated rats.

The present study was designed to evaluate the protective effect of fluted pumpkin seeds (FPS) against caffeine (CAFF) induced testicular toxicity in rats. Thirty young healthy male Wistar rats (196 ± 12 g) were randomly organized into five sets of six animals per each group: control, caffeine (CAFF; 50 mg kg-1 bw) and FPS co-treatment groups (CAFF + 50 mg FPS, CAFF + 100 mg FPS and CAFF + 200 mg FPS kg-1 bw). CAFF and FPS were administered daily and twice per week respectively by oral gavage for 40 days. CAFF treatment decreased testicular lactate dehydrogenase enzyme activity level, which was attenuated on co-administration with FPS at 50 and 100 mg kg-1 bw. Furthermore, CAFF decreased seminiferous epithelia thickness and spermatogenesis score index and increased the number of tubules with abnormal histological features, which were attenuated on co-administration with FPS at 50 mg kg-1 bw much more than at the higher doses (p < 0.05). CAFF did not affect malondialdehyde and glutathione concentrations and glutathione peroxidase (GSH-Px) activity in the testes whereas FPS co-treatment at the higher doses elevated glutathione level and GSH-Px activity and did not affect spermatogenesis score index at the highest dose (200 mg kg-1 bw). Testicular malondialdehyde concentrations remained unaffected in all FPS co-treatment groups. Overall, FPS is able to minimize the CAFF-induced testicular injury at lower than at the higher tested doses.

Increasing of LDH Specific Activity and PEPCK Level Play a Role on Activation of Gluconeogenesis Pathway in Early Onset Pre-Eeclampsia Placenta.

Recent advancement on experiment concluded that etiology of pre-eclampsia (PE) could be explained by the "two-stage" theory. The theory of which explained that pre-eclampsia occurs due to abnormalities in spiral arteries development and release of inflammatory response. Failure of spiral arteries development, the lesion of damage could be due to ischemia-reperfusion or hypoxia-reoxygenation. Hypoxia in pre-eclamptic placenta leads to metabolic change to anaerobic in glycolysis. Lactate dehydrogenase (LDH) has important role in anaerobic glycolysis that catalyzes the conversion of lactate to pyruvate during hypoxia. On the other hand, phosphoenolpyruvate carboxy kinase (PEPCK) is merely an enzyme of gluconeogenesis. This research conduct to reveal that in early onset pre-eclampsia the placenta still hypoxic and undergoes gluconeogenesis even after delivery, through metabolic enzyme of LDH and PEPCK level. This cross-sectional study compared early onset PE (< 34 weeks) with normal term placenta. We measured LDH enzyme activity using colorimetric assay and PEPCK protein using ELISA method. Result show that placental LDH specific activity was increased significantly in PE with median 2.750 (0.030 - 5.680) U/mg compared to normal term placenta 0.255 (0.032 - 1.194) U/mg (Mann-Whitney, p< 0.001). PEPCK level was significantly increased in PE 8.998 (1.737-44.914) ng/mg compared to normal term placenta 1.552 (0.741-8.832) ng/mg (Mann-Whitney, p< 0.001). We conclude that anaerobic glycolysis and gluconeogenesis pathway are increased in early onset PE placenta as adaptation mechanism to hypoxic condition.

Evaluation of a modified feeding strategy on the growth, metabolism and feeding economics of Snubnose Pompano Trachinotus blochii (Lacepede, 1801) in a recirculatory system

An experiment was conducted for 60 days to evaluate the performance of modified feeding strategy through alternate level of dietary protein in snubnose pompano, Trachinotus blochii for better nutrient utilization and to reduce feed wastage. The juveniles of snubnose pompano with an average weight of 3.0 ± 0.2 g were distributed in 12 numbers of tanks in a re-circulatory system. Four iso-caloric diets with varying level of crude protein viz., 400, 350, 300 and 250 g/kg were made and designated as P400, P350, P300 and P250. Four treatments each with triplicates were maintained based on four different feeding schedules such as regular feeding of P400 (P400/400), alternate feeding of one day P400 - next day P350 (P400/350), alternate feeding of one day P400- next day P300 (P400/300) and alternate feeding of one day P400- next day P250 (P400/250). The results showed a significant difference (P < 0.05) in specific growth rate (SGR), growth rate on metabolic body weight (GRMBW), daily growth co-efficient (DGC) and thermal growth co-efficient (TGC) among the treatments and relatively higher in treatment P400/350 followed by P400/400 and P400/300. The protein efficiency ratio (PER) and protein productive value (PPV) were higher in P400/350 treatment and lowest in P400/250. The whole body crude protein and lipid contents were found higher in P400/350 and P400/400 treatments respectively. The body indices of fish showed an increasing trend with decreased protein diet on alternate days and the maximum value was witnessed in P400/250 treatment. Higher level of serum total protein and triglycerides were recorded in P400/400 and P400/250 treatments respectively. The protease and α-amylase activities were higher in P400/350 and P400/250 treatments respectively. There was significant difference (P < 0.05) in alkaline phosphatase (ALP), acid phosphatase (ACP), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities among the treatments and showed a decreasing trend as the protein content of diet decreased in the fish fed with lower dietary protein on alternate days. Reverse trend was observed in lactate dehydrogenase (LDH) and malate dehydrogenase (MDH) enzyme activities. The P400/350 feeding schedule registered a relative feed cost savings of 4.49% in comparison with P400/400 group. The present study revealed better growth performance, protein utilization and lower feed cost in the fish of treatment P400/350, which was associated with balanced digestive and metabolic enzyme activities achieved through alternate level of higher and lower protein diet feeding.

Assessment of urine sample quality by the simultaneous measurement of urinary γ-glutamyltransferase and lactate dehydrogenase enzyme activities: possible application to unravel cheating in drugs of abuse testing.

Evaluation of the simultaneous measurement of urinary γ-glutamyltransferase (γGT) and lactate dehydrogenase (LDH) to discriminate fresh from previously frozen specimens in urine drug monitoring. Two widely available photometric tests (Siemens Healthineers Atellica) were used to determine the range of urinary γGT and LDH excretion and to study the decay in urinary enzyme activity under various storage conditions (room temperature, 4-8°C,-18°C,-80°C). From these data, cut-off values were established and evaluated in split (fresh/frozen) specimens. Both assays allow robust, reliable, and simultaneous determination of urinary γGT and LDH. In healthy subjects, the 95% reference intervals for enzyme activity in native urine were γGT: 24.4-100.4U/g Crea (creatinine) and LDH: 2.5-45.8U/g Crea. Frozen storage for at least 7days at-18°C resulted in a loss of activity to less than 50% in both enzymes. Cut-offs for frozen samples were γGT≤33.2U/g Crea and LDH≤ 8.4U/g Crea. When applied to 100 sample pairs (fresh/frozen), 86.5% (173/200) of the measurements were conclusive and the combination of concordant enzyme measurements (low γGT/low LDH or high γGT/high LDH) was able to predict the mode of storage with a sensitivity of 96.3% and a specificity of 96.7%. The additional measurements of urinary γGT and LDH can be used to detect previously frozen urine specimens. A simple protocol is proposed to provide additional information on sample quality when deceit is suspected. The procedure can be easily integrated into the standard workflow of urinary drug monitoring.

Acute and Sub-chronic Toxicological Studies of Citrus aurantium Fruit Juice in Wistar Rats

Citrus aurantium remains an excellent source of vitamins and phytochemicals. C. aurantium has been associated with lots of medicinal uses ranging from weight management to the treatment of nausea, cardiovascular diseases and cancer. This study therefore aimed at determining the acute and sub-chronic toxic effects of C. aurantium (Lemon) fruit juice in Wistar rats. The median lethal dose (LD50) for the extract was determined by Lorke’s method. A total of 20 rats of mean weight of 125 ± 5 g randomized into 4 groups of five rats each was used for the study. Groups B, C and D were administered 2ml/kg, 4ml/kg and 8ml/kg bodyweight of the extract for 3 months while group A served as the control. Random blood glucose levels of rats were monitored on monthly basis. Blood samples were collected for various biochemical assays like kidney and liver functions, serum lactate dehydrogenase enzyme activity, lipid profile, lipid peroxidation and electrolyte levels using standard diagnostic methods. Results showed a lethal dose of 70ml/kg body weight. Creatinine and urea levels were observed to decrease markedly in groups C and D against control. Significant reductions in the liver function parameters were noted for the test groups when compared with the control with a comparative significant (p&lt;0.05) increase in the activity of lactate dehydrogenase within the test groups. There was also a significant (p&lt;0.05) decrease in the levels of total cholesterol, low-density lipoprotein, very low-density lipoprotein and triglycerides within the test groups as compared with the control. The findings suggest that prolonged administration of C. aurantium does not exert any serious chronic toxicity involving tissue and organ damage. It can also be inferred that the fruit juice of C. aurantium could be employed to remedy numerous maladies that affect the body based on the lethal dose.

A novel description of the Vastus lateralis morphology of the Temminck's ground pangolin (Manis temminckii).

The pangolin (Manidae family) is the world's most trafficked animal, yet very little is known about its physiology and metabolism primarily due to its inconspicuous and solitary nature. Skeletal muscle samples from the Vastus lateralis were collected postmortem from a single female Temminck's ground pangolin (Manis temminckii). Samples were analyzed for fiber type composition, fiber size and myosin heavy chain isoform content. The oxidative and glycolytic metabolic capacity was determined through citrate synthase, 3-hydroxyacetyl co A dehydrogenase, creatine kinase, lactate dehydrogenase, phosphofructokinase and glycogen phosphorylase enzyme activities. Lastly, antioxidant capacity was determined through superoxide dismutase and catalase enzyme activities, and the total antioxidant capacity. The pangolin metabolic profile was then compared to other endurance and nonendurance mammals, in which data were standardized relative to human endurance athletes in order to provide context. Slow twitch type I fibers, rich in mitochondria were the predominant fiber type within the pangolin indicating a reliance on oxidative derived energy from fats and carbohydrates. This suggests that the pangolin has a high endurance capability when compared to other wild animals and human endurance athletes. This is the first study to investigate the skeletal muscle physiology and metabolism of any pangolin species, in an attempt to further understand this endangered animal and aid with conservation efforts.

Trehalose An Additive Solution for Platelet Concentrate to Protect Platelets from Apoptosis and Clearance during Their Storage at 4°C.

Objective Although cold storage of platelets (PLTs) could decrease the risk of bacterial growth, it could affect on the PLTs viability and hemostatic function. At cold temperatures, trehalose can be used to substitute water, inhibit the solid-liquid transition phase of the PLT membrane, and stop Glycoprotein Ibα (GPIbα) polymerization. In this study, we evaluated the potential of trehalose for reducing the negative effects of cold storage on the apoptosis and the clearance rates of PLTs after long-term storage at cold. Materials and Methods In this experimental study, PLT concentrates (PCs) were maintained for five days in the different circumstances. PLTs were subsequently counted by using an automated hematology analyzer. Also water-soluble tetrazolium salt (WST-1) assay was performed to estimate the viability of PLTs. The activity of lactate dehydrogenase enzyme (LDH) was determined by a biochemical analyzer. And human active caspase-3 levels were measured by using enzyme-linked immunosorbent assay (ELISA) method. Also, we applied flow cytometry technique. Results PLTs count and viability were higher, while LDH amount was lower in trehalose-treated PLTs when compared with two other groups (P=0.03). The highest increase in the amount of caspase-3 levels in the PLTs was observed at 4°C. However, trehalose-treated and 4°C PLTs had a lower amount of active caspase-3 in comparison with 4°C PLTs. The level of PS expression on PLTs was lower in the trehalose-treated PLTs in compared with the two other groups (P=0.03). PLTs ingestion by HepG2 cells was enhanced in the 4°C-stored PLTs. However, the ingestion rate was significantly reduced in the trehalose-treated PLTs on day 5 of storage (P=0.03). ConclusionTrehalose can moderate the effects of cold temperature on the apoptosis, viability, and the survival rate of PLTs. It also decreases the ingestion rate of refrigerated PLTs in vitro.

Polysaccharide-Protein Complex Isolated from Fruiting Bodies and Cultured Mycelia of Lingzhi or Reishi Medicinal Mushroom, Ganoderma lucidum (Agaricomycetes), Attenuates Doxorubicin-Induced Oxidative Stress and Myocardial Injury in Rats.

This study aimed to evaluate the effect of the polysaccharide-protein complex isolated from the fruiting bodies (GLFPPC) and cultured mycelia (GLMPPC) of a highly valued medicinal mushroom, Ganoderma lucidum, to alleviate doxorubicin (DOX)-induced cardiotoxicity. GLFPPC and GLMPPC were isolated from aqueous-alcoholic extracts of fruiting bodies and cultured mycelia of G. lucidum by repeated ethanol precipitation, dialysis, treatment with Sevag reagent, and freeze drying. The polysaccharide component was confirmed by assays with anthrone and phenol-sulphuric acid regents and protein moiety with Bradford reagent. The amino acid profile of protein moiety was determined by high-performance liquid chromatography analysis. DOX-induced cardiotoxicity was determined using Swiss albino mice. DOX administration caused a marked increase of creatine kinase and lactate dehydrogenase enzyme activities, indicating injury to the myocardium. The polysaccharide-protein complex downregulated cardiac injury marker enzymes, enhanced activities of endogenous antioxidants (namely, superoxide dismutase, catalase, glutathione peroxidase, and reduced glutathione levels), and significantly attenuated lipid peroxidation. The results indicated that GLFPPC and GLMPPC imparted protection against DOX-induced oxidative stress. Biochemical assays coupled with histopathological observations supported this conclusion. These experimental findings suggest that the polysaccharide-protein complex isolated from G. lucidum might be a useful therapeutic agent to ameliorate DOX-induced cardiomyopathy.

A Study to Analyze the Alkaline Phosphatase and Lactate Dehydrogenase Enzyme Activity in Gingival Crevicular Fluid During Orthodontic Tooth Movements.

Background:In orthodontic tooth development, bone turnover is described by bone testimony at locales of strain and bone resorption at areas of stress. There are metabolic disorders that may lead to tooth movement when the periodontal tissues are under mechanical stress. We needed to discover how the chemical movement of alkaline phosphatase and lactate dehydrogenase in gingival crevicular liquid (GCF) vacillated when tensions were coordinated during introductory arrangement (P1) and withdrawal (P2).Materials and Procedures:Fifty persons, ranging in age from 11 to 21 years, were enrolled in the trial, all of whom required first premolar extractions and were enduring fixed orthodontic treatment. Every subject's test and control teeth were the maxillary canine (TT) and mandibular second molar (CT). Two μL of GCF was taken from the mesial side of both TT and CT and tried for the action of ALP and LDH utilizing a spectrophotometer on the 14th day following the finish of each stage.Results:The findings were analyzed using Student's t-test. Enzyme activity changed when the teeth moved during orthodontic treatment. There was a statistically significant difference in the levels of ALP and LDH activity between P1 and P2 at TT (P = 0.005 and P = 0.001). Neither ALP nor LDH activity differed significantly between P1 and P2, with a statistical significance of 0.054 and 0.061, respectively.Conclusion:According to this research, GCF ALP and LDH activity, as well as periodontal biologic activity during orthodontic tooth movement, can be properly detected.

Inhibition of lncRNA NEAT1 protects endothelial cells against hypoxia/reoxygenation‑induced NLRP3 inflammasome activation by targeting the miR‑204/BRCC3 axis.

Cardiovascular ischemia/reperfusion (I/R) injury is primarily caused by oxygen recovery after prolonged hypoxia. Previous studies found that the long non coding RNA (lncRNA) nuclear enriched abundant transcript 1 (NEAT1) was involved in cardiovascular pathology, and that NOD-like receptor protein 3 (NLRP3) inflammasome activation-dependent pyroptosis played a key role in cardiovascular I/R injury. The present study aimed to explore the molecular mechanism of I/R pathogenesis in order to provide novel insights for potential future therapies. Cell viability and lactate dehydrogenase enzyme activity assays were used to detect cell injury after human umbilical vein endothelial cells (HUVECs) were subjected to hypoxia/reoxygenation (H/R). The expression of the NEAT1/microRNA (miR)-204/BRCA1/BRCA2-containing complex subunit 3 (BRCC3) axis was examined by reverse transcription-quantitative PCR, and the associations among genes were confirmed by luciferase reporter assays. Western blotting and ELISA were used to measure the level of NLRP3 inflammasome activation-dependent pyroptosis. The results demonstrated that NEAT1, BRCC3 expression and NLRP3 inflammasome activation-dependent pyroptosis were significantly increased in H/R-injured HUVECs, whereas silencing BRCC3 or NEAT1 attenuated H/R-induced injury and pyroptosis. NEAT1 positively regulated BRCC3 expression via competitively binding with miR-204. Moreover, NEAT1 overexpression counteracted miR-204 mimic-induced injury, BRCC3 expression and NLRP3 inflammasome activation-dependent pyroptosis. Taken together, these findings demonstrated that inhibition of lncRNA NEAT1 protects HUVECs against H/R-induced NLRP3 inflammasome activation by targeting the miR-204/BRCC3 axis.

Mesenchymal stem cells seeded onto nanofiber scaffold for myocardial regeneration

ABSTRACT Cardiac disease is the leading cause of mortality and disability worldwide. We investigated the role of undifferentiated adipose tissue-derived mesenchymal stem cells (ADMSC) alone and ADMSC seeded onto the electro-spun nanofibers (NF) for reconstructing damaged cardiac tissue in isoprenaline-induced myocardial infarction (MI) in rats. ADMSC were sorted by morphological appearance and by detection of cluster of differentiation (CD) surface antigens. The therapeutic potential of ADMSC for treating MI was evaluated by electrocardiogram (ECG), biochemical analysis, molecular genetic analysis and histological examination. Treatment of MI-challenged rats with ADMSC improved ECG findings, which were corroborated by significant decreases in serum lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) enzyme activities together with reduced serum troponin T (cTnT) and connexin 43 (Cx43) levels. MI model rats treated with ADMSC exhibited a significant increase in serum alpha sarcomeric actin (Actn) and GATA binding protein 4 (GATA4), and NK2 homeobox 5 (NKX2.5) gene expression was decreased following treatment with ADMSC. ADMSC also ameliorated damage to cardiac tissue. The effects of ADMSC seeded onto NF were superior to those of ADMSC alone. ADMSC may be useful for mitigation of MI.

Real-Time Non-Invasive and Direct Determination of Lactate Dehydrogenase Activity in Cerebral Organoids-A New Method to Characterize the Metabolism of Brain Organoids?

Organoids are a powerful tool in the quest to understand human diseases. As the developing brain is extremely inaccessible in mammals, cerebral organoids (COs) provide a unique way to investigate neural development and related disorders. The aim of this study was to utilize hyperpolarized 13C NMR to investigate the metabolism of COs in real-time, in a non-destructive manner. The enzymatic activity of lactate dehydrogenase (LDH) was determined by quantifying the rate of [1-13C]lactate production from hyperpolarized [1-13C]pyruvate. Organoid development was assessed by immunofluorescence imaging. Organoid viability was confirmed using 31P NMR spectroscopy. A total of 15 organoids collated into 3 groups with a group total weight of 20–77 mg were used in this study. Two groups were at the age of 10 weeks and one was at the age of 33 weeks. The feasibility of this approach was demonstrated in both age groups, and the LDH activity rate was found to be 1.32 ± 0.75 nmol/s (n = 3 organoid batches). These results suggest that hyperpolarized NMR can be used to characterize the metabolism of brain organoids with a total tissue wet weight of as low as 20 mg (<3 mm3) and a diameter ranging from 3 to 6 mm.

Impacts of tea tree or lemongrass essential oils supplementation on growth, immunity, carcass traits, and blood biochemical parameters of broilers reared under different stocking densities

The effects of tea tree essential oil (TTEO) and lemongrass essential oil (LGEO) with different stocking densities on the growth performance, biochemical markers, antioxidants, and immunity state of broiler chickens were studied. Birds were housed at stocking densities of 25, 30, 35, and 40 kg/m2. The treatments were, basal diet without any supplementation, the second and third groups were supplemented with 300 mg TTEO/kg feed, and 300 mg LGEO/kg feed, respectively. Results revealed that increasing stocking density from 25 to 40 kg/m2 significantly reduced body weight and daily weight gain at different ages. The phagocytic index and activity were significantly higher under the lower stocking density (25 kg/m2). Serum amyloid A (SAA), serum or liver transferrin (TRF), or C-reactive protein (CRP) were significant decreased when decreasing stocking density. Increasing stocking density from 25 to 40 kg/m2 resulted in a significant increase in the serum urea, creatinine, uric acid, lactate dehydrogenase (LDH), alanine aminotransferase (ALT), aspartate aminotransferase (AST), malondialdehyde (MDA), and catalase (CAT) levels. However, there was a significant reduction in antioxidant enzyme activity, including glutathione peroxidase (GPx) and superoxide dismutase (SOD), as stocking density increased. The supplementation of TTEO produced significantly higher body weight and daily weight gain followed by LGEO. Additionally, the mortality rates were reduced in TTEO (27.4%) and LGEO (25%) groups. TTEO or LGEO supplementation significantly improved meat constituents and cellular immunity and reduced serum total lipids, serum and meat cholesterol, and triglycerides, SAA, TRF, and CRP. For all these measured parameters, superior results were obtained when TTEO was used compared to LGEO. TTEO or LGEO supplementation also significantly reduced serum urea, creatinine, uric acid, and the enzymatic activities of LDH, ALT, AST, MDA, and CAT (but not GPx and SOD) in comparison to the control treatment. Overall, our results showed the superiority of TTEO over LGEO as a feed supplement in broiler diets. In conclusion, TTEO treatment offers a better solution for raising broiler chickens in high stocking density.

Characterisation of putative lactate synthetic pathways of Coxiella burnetii.

The zoonotic pathogen Coxiella burnetii, the causative agent of the human disease Q fever, is an ever-present danger to global public health. Investigating novel metabolic pathways necessary for C. burnetii to replicate within its unusual intracellular niche may identify new therapeutic targets. Recent studies employing stable isotope labelling established the ability of C. burnetii to synthesize lactate, despite the absence of an annotated synthetic pathway on its genome. A noncanonical lactate synthesis pathway could provide a novel anti-Coxiella target if it is essential for C. burnetii pathogenesis. In this study, two C. burnetii proteins, CBU1241 and CBU0823, were chosen for analysis based on their similarities to known lactate synthesizing enzymes. Recombinant GST-CBU1241, a putative malate dehydrogenase (MDH), did not produce measurable lactate in in vitro lactate dehydrogenase (LDH) activity assays and was confirmed to function as an MDH. Recombinant 6xHis-CBU0823, a putative NAD+-dependent malic enzyme, was shown to have both malic enzyme activity and MDH activity, however, did not produce measurable lactate in either LDH or malolactic enzyme activity assays in vitro. To examine potential lactate production by CBU0823 more directly, [13C]glucose labelling experiments compared label enrichment within metabolic pathways of a cbu0823 transposon mutant and the parent strain. No difference in lactate production was observed, but the loss of CBU0823 significantly reduced 13C-incorporation into glycolytic and TCA cycle intermediates. This disruption to central carbon metabolism did not have any apparent impact on intracellular replication within THP-1 cells. This research provides new information about the mechanism of lactate biosynthesis within C. burnetii, demonstrating that CBU1241 is not multifunctional, at least in vitro, and that CBU0823 also does not synthesize lactate. Although critical for normal central carbon metabolism of C. burnetii, loss of CBU0823 did not significantly impair replication of the bacterium inside cells.

The influence of biofilm maturation on fluoride's anticaries efficacy.

(1) To explore the influence of biofilm maturation and timing of exposure on fluoride anticaries efficacy and (2) to explore biofilm recovery post-treatment. Bovine enamel specimens were utilized in a pH cycling model (28 subgroups [n = 18]). Each subgroup received different treatments [exposure]: sodium fluoride [NaF]; stannous fluoride [SnF2]; amine fluoride [AmF]; and de-ionized water [DIW], at a specific period: early: days 1-4; middle: days 3-6; and late: days 7-10. During non-exposure periods, pH cycling included DIW instead of fluorides. Objective 1: part 1 (cycling for 4, 6, or 10days). Part 2 (cycling for 10days). Objective 2: early exposure: three sample collection time points (immediate, 3days, and 6days post-treatment); middle exposure: two sample collection time points (immediate, 4days post-treatment). The enamel and biofilm were analyzed ([surface microhardness; mineral loss; lesion depth]; [lactate dehydrogenase enzyme activity; exopolysaccharide amount; viability]). Data were analyzed using ANOVA (p = 0.05). Objective 1: Early exposure to fluorides produced protective effects against lesion progression in surface microhardness and mineral loss, but not for lesion depth. Objective 2: Early exposure slowed the demineralization process. SnF2 and AmF were superior to NaF in reducing LDH and EPS values, regardless of exposure time. They also prevented biofilm recovery. Earlier exposure to SnF2 and AmF may result in less tolerant biofilm. Early fluoride treatment may produce a protective effect against demineralization. SnF2 and AmF may be the choice to treat older biofilm and prevent biofilm recovery. The study provides an understanding of biofilm-fluoride interaction with mature biofilm (e.g., hard-to-reach areas, orthodontic patients) and fluoride's sustainable effect hours/days after brushing.

Correlation of Lactate Dehydrogenase (LDH) activity, D-Dimer and Severity in a sample of COVID-19 patients in Iraq

Subject: The coronavirus pandemic, which began in 2019, is a global outbreak of coronavirus disease. The World Health Organization (WHO) also classified Severe Acute Respiratory Syndrome (SARS) as a new coronavirus (SARS-CoV-2). Wuhan, China, was the first place in December 2019 when the pandemic was found. LDH levels were related to a six-fold increase in the risk of developing severe COVID-19 disease. The objective of the study: Estimation of enzyme activity of Lactate Dehydrogenase enzyme (LDH) and d-dimer in patients with COVID-19 to estimate the behavior of enzyme during the period of the disease. Correlation of enzyme activity Lactate Dehydrogenase (LDH)with d-dimer. More crucially, elevated LDH was related to a 16-fold increase in the risk of death. Abnormal coagulation function, including elevated D-dimer, has been demonstrated to be more common in deceased patients with COVID-19, and increasing odds of in-hospital death were associated with D-dimer greater than 1 μg/ml. The current study aimed to investigate serum levels of LDH and d- dimer in the severity of COVID-19 patients and to clarify the correlation of these parameters with disease severity Material and method: This study included (48) patients with COVID-19 in severity (confirmed by polymerase chain reaction). COVID-19 patients showed a significant elevation in the levels of all parameters, we concluded a proportional correlation between the aforementioned parameters and COVID-19 suggesting the uses of these tests to the diagnosis of severe cases. Results: This study showed a high correlation between the activity of lactate dehydrogenase (LDH) and disease progression and severity in patients with COVID-19. Also, the study showed a high correlation between D-dimer and disease progression and severity in patients with COVI19. Conclusion: There is a correlation between lactate dehydrogenase (LDH)level and D- dimer level later on during disease Progression. But not and the time of hospitalization.