scholarly journals VDR Pathway-Specific Dose-Related Effects of 1,25(OH)2D3 on Salmonella typhimurium-induced Mouse Colitis

2020 ◽  
Author(s):  
Fubin Qiu ◽  
Lijuan Zhang ◽  
Chunyan Li ◽  
Ying Zhang ◽  
Xiaoyan Zhang
Keyword(s):  
Mrna Expression ◽  
Current Data ◽  
Expression Level ◽  
Control Group ◽  
High Dose ◽  
Mrna Expression Level ◽  
Significant Difference ◽  
Dose Limits ◽  
Group A ◽  
Cervical Dislocation

Abstract Background: Whether and how 1,25(OH)2D3 supplementation influences VDRs in experimental mice with colitis remains to be seen. To explore the effect of 1,25(OH)2D3 on S. typhimurium colitis through the VDR pathway and to discover the role of VDR in its action. Methods: We established a mouse UC model induced by S. typhimurium. After streptococcal typhus infection, the mice were fasted for 12 hours. Blood was collected by the eyeball extraction method, and then sacrificed by cervical dislocation, specimens were collected for corresponding indicators. Results: Mice exposed to S. typhimurium infection developed signs of acute colitis. After HE staining were performed on the diseased colons from the mice. high dose VD supplementation, the pathological colonic damage did not improve in the mice, and there was no statistical difference between the groups with VD deficiency (P>0.05). VDR expression in the UC group treated with Salmonella was higher than that in the control group, a statistically significant difference (P<0.01). Compared with the VDD+UC group, VDR expression rose in both the LVDS+UC group and the HVDS+UC group, with VDR protein expression being highest after high dose VD supplementation (P<0.01). Compared with the control and the UC groups, the VDR mRNA expression level in the VDD+UC group was significantly higher, and the colon VDR mRNA expression level decreased after active VD supplementation (Fig.7C). Conclusions: Our data suggest the need for defining the accurate 1,25(OH)2D3 dose limits that induce an anti-inflammatory effect as current data indicate that higher doses would produce an inflammatory response.

195 EFFECTS OF GSK3 INHIBITOR ON THE PLURIPOTENCY MAINTENANCE OF BUFFALO EMBRYONIC STEM-CELL-LIKE CELLS

2014 ◽  
Vol 26 (1) ◽  
pp. 212 ◽  
Author(s):  
F. Lu ◽  
Y. Lao ◽  
H. Sun ◽  
C. Lei ◽  
Y. Deng ◽  
...  
Keyword(s):  
Stem Cell ◽  
Embryonic Stem Cell ◽  
Mrna Expression ◽  
Colony Formation ◽  
Embryonic Stem ◽  
Signalling Pathway ◽  
Expression Level ◽  
Control Group ◽  
Mrna Expression Level ◽  
Significant Difference

In this study, to explore the effects and mechanism of Wnt/β-catenin signalling pathway on the maintenance of pluripotency of buffalo embryonic stem-cell-like cells (buffalo ESC-like cells), the GSK3 inhibitors BIO and CHIR99021 were added throughout the experiment – i.e. from buffalo inner cell mass (ICM) culture to ESC-like line generation. The buffalo ICM were respectively cultured in the medium containing 0.5 μg mL–1 BIO and 5 mmol L–1 CHIR99021. The percentage of ICMs attachment and primary colony formation were observed, and found that there was no significant difference in the ICMs attachment rate among of the BIO, CHIR99021, and the control groups (91.18% and 92.98% v. 94.59%; P > 0.05). Treating ICMs with CHIR99021 resulted in more primary colony formation rate compared with the control group (77.71% v. 55.41%; P < 0.05). The proliferation rate of primary colonies of buffalo ESC-like cells was detected by bromodeoxyuridine immunofluorescence techniques. The results show that the proliferation rate of primary colonies in the group of buffalo ESC-like cells treated with CHIR99021 was significantly higher than that of the control group on Day 1, Day 3, Day 4, and Day 5 (P < 0.05), and it was also evidently higher than that of control group only on Day 1 (P < 0.05) in the group of BIO, but there was no significant difference in other days (P > 0.05). The mRNA expression level of proliferation marker PCNA of ESC-like cells was significantly up-regulated in both CHIR99021 and BIO treatment groups (P < 0.05), however, treating buffalo ESC-like cells with CHIR99021 significantly up-regulated the expression of pluripotent gene Oct4 and Sox2 (P < 0.05), but had no effect on pluripotent gene Nanog expression (P > 0.05). Oct4 expression was significantly increased (P < 0.05), and the expression of Sox2 and Nanog were significantly decreased (P < 0.05) in the group of BIO treatment. Furthermore, the relative protein level of β-catenin (the downstream effector of Wnt/β-catenin signalling pathway) and the mRNA expression level of c-Myc (the downstream target gene of Wnt/β-catenin signalling pathway) were significantly increased when buffalo ESC-like cells respectively treated with CHIR99021 and BIO (P < 0.05). In conclusion, treating buffalo ESC-like cells with GSK3 inhibitors CHIR99021 can promote proliferation of buffalo ESC-like cells, maintain their undifferentiated state, and up-regulate the expression levels of β-Catenin and c-Myc in buffalo ESC-like cells. These results indicate that Wnt/β-catenin signalling pathway plays an important role in regulation of self-renewal of buffalo ESC-like cells. This work was funded by the China High Technology Development Program (2011AA100607), China Natural Science Foundation (31072033), and Guangxi Science Foundation (2012GXNSFFA060004).

scholarly journals PSIV-B-38 Late-Breaking: Effect of Glutamate (Glu) on Developmental Block and the MZT Relative Genes Expression in Mouse embryo during in Vitro Culture

2019 ◽  
Vol 97 (Supplement_3) ◽  
pp. 330-331
Author(s):  
Yu Liu ◽  
An Gang Lou ◽  
Shuo Yang ◽  
Zhong Shu Li ◽  
Nan-Zhu Fang
Keyword(s):  
In Vitro Culture ◽  
Mrna Expression ◽  
Marker Gene ◽  
Treated Group ◽  
Mouse Embryos ◽  
Expression Level ◽  
Mrna Expression Level ◽  
Significant Difference ◽  
Developmental Block

Abstract The risk of developmental block in mammal’s embryos is high during in vitro as compare to in vivo environment because the in vitro embryo-culture systems are suboptimal. During in vitro-culture the balance between ROS production and elimination is disturbed and may lead to 2-cell block in mouse embryos [1]. In the current study, we investigated the effects of Glu as anti-developmental block during IVC on ZGA and MZT on mouse embryos. The mouse embryos were divided into control and different level of Glu treated group. The cleavage rate was determined, the ROS and GSH level was investigated using DCHF-DA and CMF2HC respectively. The mRNA expression level of ZGA marker gene such as Eif-1α, Muerv l, Zscan4d and Hsp70.1 was analyzed among the groups using RT-PCR. The transition rate from 2-cell to 4-cell was significantly higher in 6mmol/L Glu treated group as compare to control and others treated groups. No significant difference was recorded in the level of ROS and GSH during MZT stage among the different groups. The mRNA expression level of ZGA marker gene was significantly increased at middle and late stage in 6mmol/L Glu treated group as compare to control and others treated groups. In conclusion, this study shows that the concentration of 6mmol/L Glu could maintain the dynamic balance of GSH and ROS, increase the expression of ZGA marker gene and maintain its high expression pattern of time series, directly participate in the ZGA activated process; ultimately reduce the risk of developmental block to ensure the successful completion of MZT. Reference [1] Lee MT, Bonneau AR, Giraldez AJ.Zygotic Genome Activation during the Maternal-to-Zygotic Transition. Annual Rev Cell Dev Biol [J], 2014, 30:581–613.

scholarly journals 1Hz and 100mT Electromagnetic Field Induces Apoptosis in Breast Cancer Cells Through Up-Regulation of P38 and P21

2020 ◽  
Vol 4 (1) ◽  
pp. 23-29
Author(s):  
Akram Sajadian ◽  
Elham Razmpoosh ◽  
Farshid Alaeddini ◽  
Maryam Bassiri
Keyword(s):  
Breast Cancer ◽  
Electromagnetic Field ◽  
Mrna Expression ◽  
Cell Line ◽  
Cancer Cell Line ◽  
Underlying Mechanism ◽  
Expression Level ◽  
Control Group ◽  
Mrna Expression Level ◽  
Sham Exposure

Introduction: Breast cancer is the most common cause of cancer-related death among women. Recently, extremely low-frequency electromagnetic field (ELF-EMF) has been proposed as a new interfering agent with future therapeutic potentials. Many studies have revealed that cellular processes such as apoptosis in breast cancer are affected by ELF-EMFs. However, more researches are needed to clarify the underlying mechanism of action for these fields. In this study, the apoptotic effect of ELF-EMF on the MC4L2 cell line was examined and the mRNA expression level of the P21 and P38 genes were further investigated. Methods: A triple-positive mouse breast cancer cell line (MC4L2) was purchased from the Genetic Resource Center (Iran). This study was performed on two groups of ELF-EMF exposure (100mT/1 Hz for 5 days, 120 min each day) and sham exposure. Cell viability and apoptosis rate of both the exposure and sham exposure groups weredetermined by flow cytometry. Alterations in the P21 and P38 mRNAs expression levelswere investigated; using real-time PCR. Results: ELF-EMF exposure induced 30% apoptosis in MC4L2 cells compared with the control group. The mRNA expression level of P38 and P21 was significantly increased after ELF-EMF exposure compared to the control group. Conclusions: ELF-EMF induces apoptosis in the MC4L2 triple-positive cell line. Furthermore, this exposure affects important gene expression involved in the cell cycle. Our data propose that ELF-EMF in a specific time, intensity, and frequency could be beneficial for breast cancer treatment. However, more studies are required to confirm our findings.

scholarly journals Effects of Dietary Zinc Sources on Growth Performance and Gut Health of Weaned Piglets

2021 ◽  
Vol 12 ◽  
Author(s):  
Hui Diao ◽  
Jiayou Yan ◽  
Shuwei Li ◽  
Shengyao Kuang ◽  
Xiaolan Wei ◽  
...  
Keyword(s):  
Growth Performance ◽  
Mrna Expression ◽  
Dietary Supplementation ◽  
Basal Diet ◽  
Expression Level ◽  
Control Group ◽  
Mrna Expression Level ◽  
Gut Health ◽  
Weaned Piglets ◽  
Zinc Sources

The present study aimed to investigate the effects of dietary zinc sources on the growth performance and gut health of weaned piglets. In total, 96 Duroc × Landrace × Yorkshire (DLY) weaned piglets with an initial average body weight of 8.81±0.42kg were divided into four groups, with six replicates per treatment and four pigs per replicate. The dietary treatment groups were as follows: (1) control group, basal diet; (2) zinc sulphate (ZnSO4) group, basal diet +100mg/kg ZnSO4; (3) glycine zinc (Gly-Zn) group, basal diet +100mg/kg Gly-Zn and (4) zinc lactate group, and basal diet +100mg/kg zinc lactate. The whole trial lasted for 28days. Decreased F/G was noted in the Gly-Zn and zinc lactate groups (p&lt;0.05). The zinc lactate group had a lower diarrhea rate than the control group (p&lt;0.05). Moreover, the ZnSO4, Gly-Zn, and zinc lactate groups had significantly higher apparent total tract digestibility of dry matter (DM), crude protein (CP), ether extract (EE), crude ash, and zinc than the control group (p&lt;0.05). The Gly-Zn and zinc lactate groups had higher jejunal villus height and a higher villus height:crypt depth ratio than the control group (p&lt;0.05). In addition, the ZnSO4, Gly-Zn and zinc lactate groups had a significantly lower mRNA expression level of jejunal ZRT/IRT-like protein 4 (ZIP4) and higher mRNA expression level of jejunal interleukin-1β (IL-1β) than the control group (p&lt;0.05). The mRNA expression level of jejunal zinc transporter 2 (ZNT2) was higher and that of jejunal Bcl-2-associated X protein (Bax) was lower in the Gly-Zn and zinc lactate groups than in the control group (p&lt;0.05). Moreover, the zinc lactate group had a higher count of Lactobacillus spp. in the cecal digesta and higher mRNA expression levels of jejunal occludin and mucin 2 (MUC2) than the control group (p&lt;0.05). In conclusion, dietary supplementation with 100mg/kg ZnSO4, Gly-Zn, or zinc lactate could improve the growth performance and gut barrier function of weaned piglets. Dietary supplementation with organic zinc, particularly zinc lactate, had the best effect.

Adoption of Biomedical Ceramic iRoot BP in the Treatment of Localized Pulpitis in Children

2022 ◽  
Vol 12 (1) ◽  
pp. 174-182
Author(s):  
Baoying Peng ◽  
Na Feng ◽  
Junyan Tan
Keyword(s):  
Osteogenic Differentiation ◽  
Mrna Expression ◽  
Signaling Pathway ◽  
Clinical Efficacy ◽  
Mapk Signaling ◽  
Expression Level ◽  
Mrna Expression Level ◽  
Group A ◽  
Group B ◽  
Better Than

To explore the clinical efficacy of biomedical ceramic iRoot BP in the treatment of localized acute pulpitis in children, and the effect of iRoot BP on proliferation and osteogenic differentiation of human dental pulp stem cells (hDPSCs), 72 localized acute pulpitis children admitted to our hospital from September 2018 to September 2019 were selected and divided into group A (treated with MTA pulp capping material) and group B (treated with iRoot BP material), and the clinical effect, pain degree, and adverse reactions (ADR) rate were compared. The effects of iRoot BP on hDPSCs proliferation and osteogenic differentiation were analyzed; the proliferative activity of cells in iRoot BP group, MTA group, and control group (C group) were measured by cholecystokinin-8 (CCK-8) assay, the ability of cell mineralized nodular formation was observed via alizarin red staining; and quantitative reverse transcription PCR (qRT-PCR) andWestern blot were adopted to determine the expression of osteogenic related genes of hDPSCs and key proteins of mitogen-activated protein kinase (MAPK) signaling pathway. After 1 week of treatment, the clinical efficacy of group B was more favorable in contrast with group A (P < 0.05); the pain of children in group B was notably better in contrast with group A, and incidence of ADR was notably lower in contrast with group A (P < 0.05). 5.0 mg/mL, 10.0 mg/mL, and 30 mg/mL iRoot BP or MTA could improve cell proliferation activity (P < 0.01); the effect of iRoot BP on proliferation of hDPSCs was greater in contrast with MTA (P < 0.05); and the integral optical density (IOD) value of iRoot BP group was notably higher in contrast with MTA group (P < 0.01). The mRNA expression levels of collagen-I (COL-I), bone sialoprotein (BSP), and osteocalcin (OC) in MTA group and iRoot BP group were notably higher in contrast with C group (P < 0.01); the COL-I mRNA expression level of iRoot BP group was notably higher in contrast with MTA group (P < 0.01); the mRNA expression level of BSP in MTA group was notably higher in contrast with iRoot BP group (P < 0.01); the relative protein expression levels of phosphorylated ERK (p-ERK) and phospho-Jun N-terminal kinase (p-JNK) in MTA group and iRoot BP group were notably higher in contrast with C group (P < 0.01); and the relative expression level of p-ERK protein in iRoot BP group was higher in contrast with MTA group (P < 0.05). These results indicated that the clinical efficacy of biomedical ceramic iRoot BP was better than MTA in the preservation of live pulpitis in children, and the patients treated with iRoot BP had better pain recovery effect and lower risk of ADR. The effect of iRoot BP on the proliferation and mineralization of hDPSCs was better than that of MTA, and it may promote the osteogenic differentiation of hDPSCs by activating MAPK signaling pathway and regulating gene expression of COL-I, BSP, and OC.

Amifostine Can Reduce Oral Mucositis after High-Dose Melphalan Conditioning in Patients with Multiple Myeloma: A Retrospective Study.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 5022-5022
Author(s):  
Antoine Italiano ◽  
Emmanuel Chamorey ◽  
Cyril Foa ◽  
Frédéric Peyrade ◽  
Josiane Otto ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Oral Mucositis ◽  
Anticancer Agents ◽  
Systemic Infection ◽  
Control Group ◽  
High Dose ◽  
Significant Difference ◽  
Group A ◽  
Group B ◽  
High Dose Melphalan

Abstract Autologous peripheral blood stem cell (PBSC)- supported high-dose melphalan (HDM) is now considered standard therapy in the treatment of multiple myeloma, at least for patients under 65 years. Oral mucositis is a frequent non-hematological complication which causes severe pain, interferes with patient nutrition and can lead to systemic infection. Amifostine (WR-2721; Ethyol), a phosphoaminothiol, is a prodrug that protects a broad range of normal tissues from the cytotoxic damage induced by anticancer agents. We retrospectively compared two groups of patients with stage II/III previously untreated multiple myeloma who received between April 96 and May 2004 an induction chemotherapy with 3 or 4 cycles of VAD (vincristine, adriamycin, dexamethasone) followed by HDM (200 mg/m²) and autologous PBSC transplantation. These two groups either received (group A, n = 10 ) or did not receive (group B, n= 32) amifostine (740 mg/m²) before HDM. The occurrence of grade 3/4 oral mucositis was significantly decreased in group A in comparison to group B (10% versus 53%, p =0.023) with no difference for the time to mucosal recovery. Supportive care differed between the two groups: only 2 patients (20%) needed opioid treatment in group A versus 22 patients (69%) in group B ( p=0.005) and 1 patient (10%) required parenteral nutrition in group A compared to 16 patients (50%) in group B (p= 0.015). The occurrence of severe infectious complications did not differ between the two groups (0% versus 12,5%, p= 0.56). Amifostine did not affect haematological recovery, the median time to granulocyte recovery to > 500/μl was similar in the two groups (8,4 days versus 9,9 days, p=0.22). Moreover, there is no statistically significant difference between the amifostine and control group for the disease response. The tolerability of amifostine was excellent and no adverse effects were reported. This study suggest that amifostine can reduce mucosal damage associated with high dose melphalan-based therapy, reducing, as a consequence, the necessity of nutrition and analgesic support, without compromising therapeutic benefit. Obviously, these interesting results have to be confirmed by larger randomised trials.

scholarly journals Role of FRG1 in predicting the overall survivability in cancers using multivariate based optimal model

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Rehan Khan ◽  
Ananya Palo ◽  
Manjusha Dixit
Keyword(s):  
Mrna Expression ◽  
Cancer Patients ◽  
Risk Score ◽  
Cox Regression ◽  
Expression Level ◽  
Risk Scores ◽  
Mrna Expression Level ◽  
Liver Cancers ◽  
Significant Difference

AbstractFRG1 has a role in tumorigenesis and angiogenesis. Our preliminary analysis showed that FRG1 mRNA expression is associated with overall survival (OS) in certain cancers, but the effect varies. In cervix and gastric cancers, we found a clear difference in the OS between the low and high FRG1 mRNA expression groups, but the difference was not prominent in breast, lung, and liver cancers. We hypothesized that FRG1 expression level could affect the functionality of the correlated genes or vice versa, which might mask the effect of a single gene on the OS analysis in cancer patients. We used the multivariate Cox regression, risk score, and Kaplan Meier analyses to determine OS in a multigene model. STRING, Cytoscape, HIPPIE, Gene Ontology, and DAVID (KEGG) were used to deduce FRG1 associated pathways. In breast, lung, and liver cancers, we found a distinct difference in the OS between the low and high FRG1 mRNA expression groups in the multigene model, suggesting an independent role of FRG1 in survival. Risk scores were calculated based upon regression coefficients in the multigene model. Low and high-risk score groups showed a significant difference in the FRG1 mRNA expression level and OS. HPF1, RPL34, and EXOSC9 were the most common genes present in FRG1 associated pathways across the cancer types. Validation of the effect of FRG1 mRNA expression level on these genes by qRT-PCR supports that FRG1 might be an upstream regulator of their expression. These genes may have multiple regulators, which also affect their expression, leading to the masking effect in the survival analysis. In conclusion, our study highlights the role of FRG1 in the survivability of cancer patients in tissue-specific manner and the use of multigene models in prognosis.

scholarly journals The parp-1 and bax genes as potential targets for treatment of the heart functioning impairments induced by type 1 diabetes mellitus

2021 ◽  
Vol 55 (2) ◽  
pp. 61-71
Author(s):  
Tamara Kuchmerovska ◽  
Mykhailo Guzyk ◽  
Tetiana Tykhonenko ◽  
Lesya Yanitska ◽  
Irina Pryvrotska ◽  
...  
Keyword(s):  
Diabetes Mellitus ◽  
Vitamin D ◽  
Mrna Expression ◽  
Diabetic Rats ◽  
Expression Level ◽  
Total Serum ◽  
Control Group ◽  
Mrna Levels ◽  
Mrna Expression Level ◽  
Parp 1

Abstract Objectives. The present study was designed to assess whether apoptosis-related genes as parp-1 and bax could be targets for treatment of diabetes mellitus and whether vitamin D may exert beneficial effects. Methods. Vitamin D3 treatment for 4 weeks, starting after 4 weeks of the diabetes duration. The expression of parp-1 and bax genes was estimated on mRNA levels using real time quantitative polymerase chain reaction. Results. After 8 weeks, diabetic rats had weight loss, while blood glucose was increased about 4.9-fold compared to control group. Vitamin D3 administration to diabetic animals had no effect on these parameters. It was found that total serum alkaline phosphatase activity was significantly elevated in diabetic rats as compared to control animals and was restored by vitamin D3. Diabetes was accompanied by reduction of nicotinamidadenindinucleotide, a substrate of poly-ADP-ribosylation, level by 31.7% as compared to control rats, which was not reversed in response to vitamin D3 treatment. In diabetic hearts, the mRNA expression level of parp-1 gene was 2.8-fold higher compared to control rats and partially decreased by vitamin D3 treatment. Less significant alterations were observed in diabetic hearts for the mRNA expression level of bax gene that was 2.0-fold higher compared to control animals and vitamin D3 normalized it. These results indicate that cardiomyocytes have a tendency to apoptosis. Conclusions. The findings suggest that investigated genes can be targets at the transcriptional level for vitamin D action that may be contributed to the improving metabolic/signaling pathways induced by diabetes mellitus.

scholarly journals Analysis of DNA methylation level and mRNA expression of Transient Receptor Ankyrin Member 1 (TRPA1) in endometriosis-associated pain

2021 ◽  
pp. 1-10
Author(s):  
Ocktariyana ◽  
Nurul Hikmawati ◽  
Andon Hestiantoro ◽  
Raden Muharam ◽  
Muhammad Luky Marwali ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Mrna Expression ◽  
Methylation Level ◽  
Rating Scale ◽  
Expression Level ◽  
Pain Level ◽  
Mrna Expression Level ◽  
Significant Difference ◽  
Peritoneal Endometriosis ◽  
Transient Receptor

Transient Receptor Ankyrin Member 1 (TRPA1) is an ion channel family protein that regulates pain sensation through sensory neurons' activity. This study's purpose to analyzes the DNA methylation and mRNA expression level of the TRPA1 gene in endometriosis and its correlation with pain level. Twenty samples of peritoneal endometriosis and endometrial samples were obtained from women with endometriosis, which was subsequently compared to 20 endometrial samples of women without endometriosis. The DNA methylation level of TRPA1 was analyzed using Methylation-specific PCR (MS-PCR) and ImageJ software, while the mRNA expression of TRPA1 was analyzed using qRT-PCR. Furthermore, the pain level was measured using the numeric rating scale (NRS) by interviewing all the women. This study showed that there was a significant difference in the mRNA expression of TRPA1 in peritoneal endometriosis. The TRPA1 was unmethylated in both peritoneal and endometrial samples in endometriosis. However, DNA Methylation level of TRPA1 in peritoneal and endometrial of endometriosis compared to normal endometrial were no significant difference. Additionally, there was no correlation between DNA methylation level and mRNA expression level of TRPA1 in all samples, along with the endometriosis-associated pain.

scholarly journals Pyruvate Kinase M2 Regulates Hif-1alpha Activity in Myeloid Dendritic Cells from Patients with Severe Aplastic Anemia

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 1109-1109
Author(s):  
Xiaowei Liang ◽  
Lijie Zeng ◽  
Xiaoyu Zhao ◽  
Chunyan Liu ◽  
Zonghong Shao ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Dendritic Cells ◽  
Aplastic Anemia ◽  
Mrna Expression ◽  
Immune Cells ◽  
Expression Level ◽  
Control Group ◽  
Mrna Expression Level ◽  
Myeloid Dendritic Cells ◽  
Normal Controls

Abstract Hypoxia-inducible factor 1 (HIF-1) is a nuclear protein with transcriptional activity. HIF-1 can be activated by immune cells exposed to hypoxia and regulate glycolysis. In this regulation, pyruvate kinase M2 (PKM2), a key enzyme in the cell glycolytic pathway, is an important regulatory site. Our previous studies suggest that PKM expression is increased in myeloid dendritic cells (mDCs) in SAA patients. Therefore, we investigated the expression level of HIF-1α in mDCs and its interaction with PKM2 in SAA patients. The HIF-1α expression of mRNA and protein on mDCs in SAA untreated patients was significantly higher than that in SAA remission patients and normal controls. In the SAA patients, the HIF-1α expression on mDCs was positively correlated with the numbers of mDCs (p &lt; 0.01), CD80+mDC/mDC (r = 0.689, p &lt; 0.01), and CD86+mDC/mDC in PB (p &lt; 0.05). In SAA patients, the HIF-1α expression on mDCs was negatively correlated with the CD4+T/CD8+T ratio in peripheral blood (PB) (p &lt; 0.05), the percentage of granulocytoid and erythroid cells in bone marrow (p &lt; 0.05), the WBC count in PB (p &lt; 0.05), ANC in PB (p &lt; 0.05), and the percentage of Rets in the PB (p &lt; 0.05); was positively correlated with the percentage of lymphoid cells in bone marrow(p &lt; 0.05); and was not statistically correlated with the megakaryocyte number in bone marrow, absolute PBL count, HGB in PB, absolute Ret count in PB, or PLT count in PB. In the correlation analysis we observed that there was a positively correlation between HIF-1α and PKM2 expression on mDCs in SAA patients (p &lt; 0.001). To evaluate whether there is a mutual adjustment relationship between HIF-1α and PKM2, we successfully reduced PKM2 gene expression in this cell population via siRNA transfection. This process resulted in significantly lower levels of PKM2 protein expression relative to cells transfected with siControl which were evaluated by western blotting. We observed that the relative expression levels of HIF-1α mRNA in mDCs transfected with PKM2 siRNA was lower than siControl group（P&lt; 0.01）. Conclusions  In this study, we found that untreated patients with SAA had higher HIF-1α expression on mDCs compared with recovering SAA patients and normal controls. The expression of HIF-1α was correlated with the number and function in mDCs and the severity of pancytopenia of SAA. The results indicated that the mRNA expression level of HIF-1α in mDC of SAA patients at the onset was significantly higher than that of the remission group and the control group, and was significantly positively correlated with the expression of PKM2. The mRNA expression level of HIF-1α in mDC of SAA patients at the onset was significantly higher than that of the remission group and the control group, and was significantly positively correlated with the expression of PKM2. The above experimental results confirmed that HIF-1α plays an important role in the abnormal immune response in patients with severe aplastic anemia, mainly by regulating the activity of PKM2 and thereby affecting the energy metabolism in immune cells. Disclosures No relevant conflicts of interest to declare.

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