Perspectives on RNA Interference in Immunopharmacology and Immunotherapy

RNA interference (RNAi), mediated by short interfering RNA (siRNA), vector-derived short hairpin RNA (shRNA) and microRNA (miRNA), brings about revolutionary features to basic biomedical research and clinical application. New drugs based on RNAi have been developed for therapeutic applications. The family of RNAi molecules are efficient agents to modulate mammalian immune system, and many studies reported that these molecules could manipulate immune defence, surveillance and homeostasis. Both perfect match of siRNA/shRNA and non-perfect match of miRNA could be beneficial for designing RNAi-based drugs for treatment of tumour and viral infection. This chapter provides a view to control or utilize the immune regulation of various small RNAs that should help researchers to understand the successful clinical application of RNAi

Functional characterization of the mitochondrial 12S rRNA C1494T mutation associated with aminoglycoside-induced and non-syndromic hearing loss

In this study, we report the biochemical characterization of the deafness-associated mitochondrial 12S rRNA C1494T mutation using 27 cybrid cell lines constructed by transferring mitochondria from 9 lymphoblastoid cell lines derived from a Chinese family into human mitochondrial DNA (mtDNA)-less (ρ°) cells. Six cybrids derived from two asymptomatic members, and nine cybrids derived from three symptomatic members of the Chinese family carrying the C1494T mutation exhibited ∼38 and 43% decrease in the rate of mitochondrial protein labeling, respectively, compared with twelve cybrids derived from four Chinese control individuals. These defects are apparently a primary contributor to significant reductions in the rate of overall respiratory capacity or the rate of malate/glutamate promoted respiration, or succinate/G3P-promoted respiration, or TMPD/ascorbate-promoted respiration in mutant cybrid cell lines derived from either symptomatic or asymptomatic individuals. Furthermore, the very significant/nearly identical increase in the ratio of doubling times in DMDM medium in the presence/absence of high concentration of paromomycin was observed in symptomatic or asymptomatic cybrid cell lines carrying the C1494T mutation as compared with the average rate in control cell lines. These observations provide the direct biochemical evidences that the C1494T mutation is a pathogenic mtDNA mutation associated with aminoglycoside-induced and non-syndromic hearing loss. In addition, these data provide the first biochemical evidence that nuclear background plays a critical role in the phenotypic manifestation of non-syndromic hearing loss and aminoglycoside toxicity associated with the C1494T mutation

Folate monoglutamate in cereal grains: Evaluation of extraction techniques and determination by LC-MS/MS

Folates are essential micronutrients for human health. To determine the total folate content, the extraction and quantification of seven monoglutamate folate derivatives in cereals (maize, rice, and wheat) were optimised and validated in this study. Di-enzyme treatment with α-amylase and rat conjugase was proved ideal for folate extraction from the cereal grains. The quantification method by liquid chromatography-tandem mass spectrometry was validated based on its matrix effect, linearity, sensitivity, recovery, inter-day and intra-day precision. The limits of detection and quantification of folate derivatives ranged from 0.03–0.88 and 0.1–1.0 μg/100 g among the three cereal samples. The absolute recoveries of most folate derivatives were 72–96 % for these cereal samples, with the exception of dihydrofolate, tetrahydrofolate, and folic acid (44–65 %). The inter-day and intra-day precisions were < 12 % for the three cereals. Analysis of folate content and composition in several cereal grains showed that the total folate levels were approximately 26−37 μg/100 g, with 5-methyl-tetrahydrofolate and 5-methyl-tetrahydrofolate as the dominant. MeFox, an oxidation product of 5-methyltetrahydrofolate, was detected at concentrations 20–39-fold higher than those of total folates in rice and wheat grains. This validated method is an efficient approach for folate determination in cereal crops

hElp3 Directly Modulates the Expression of HSP70 Gene in HeLa Cells via HAT Activity

Human Elongator complex, which plays a key role in transcript elongation in vitro assay, is incredibly similar in either components or function to its yeast counterpart. However, there are only a few studies focusing on its target gene characterization in vivo. We studied the effect of down-regulation of the human elongation protein 3 (hELP3) on the expression of HSP70 through antisense strategy. Transfecting antisense plasmid p1107 into HeLa cells highly suppressed hELP3 expression, and substantially reduced expression of HSP70 mRNA and protein. Furthermore, chromatin immunoprecipitation assay (ChIP Assay) revealed that hElp3 participates in the transcription elongation of HSPA1A in HeLa cells. Finally, complementation and ChIP Assay in yeast showed that hElp3 can not only complement the growth and slow activation of HSP70 (SSA3) gene transcription, but also directly regulates the transcription of SSA3. On the contrary, these functions are lost when the HAT domain is deleted from hElp3. These data suggest that hElp3 can regulate the transcription of HSP70 gene, and the HAT domain of hElp3 is essential for this function. These findings now provide novel insights and evidence of the functions of hELP3 in human cells

Effects of oat (Avena sativa L.) hay diet supplementation on the intestinal microbiome and metabolome of Small-tail Han sheep

Supplementation of the sheep diet with oats (Avena sativa L.) improves animal growth and meat quality, however effects on intestinal microbes and their metabolites was not clear. This study aimed to establish the effect of dietary oat supplementation on rumen and colonic microbial abundance and explore the relationship with subsequent changes in digesta metabolites. Twenty Small-tail Han sheep were randomly assigned to a diet containing 30 g/100 g of maize straw (Control) or oat hay (Oat). After 90-days on experimental diets, rumen and colon digesta were collected and microbial diversity was determined by 16S rRNA gene Illumina NovaSeq sequencing and metabolomics was conducted using Ultra-high performance liquid chromatography Q-Exactive mass spectrometry (UHPLC-QE-MS). Compared to Control group, oat hay increased the abundance of Bacteroidetes and Fibrobacteres as well as known short-chain fatty acid (SCFA) producers Prevotellaceae, Ruminococcaceae and Fibrobacteraceae in rumen (p &lt; 0.05). In rumen digesta, the Oat group showed had higher levels of (3Z,6Z)-3,6-nonadienal, Limonene-1,2-epoxide, P-tolualdehyde, and Salicylaldehyde compared to Control (p &lt; 0.05) and these metabolites were positively correlated with the abundance of cecal Prevotellaceae NK3B31. In conclusion, supplementation of the sheep diet with oat hay improved desirable microbes and metabolites in the rumen, providing insight into mechanisms whereby meat quality can be improved by oat hay supplementation

Identification of Actual Irrigated Areas in Tropical Regions Based on Remote Sensing Evapotranspiration

Amidst global climate change and unsustainable human exploitation of water resources, water has emerged as a critical factor constraining global agricultural food production and ecological environments. Particularly in agricultural powerhouses like China, irrigation water accounts for a significant portion of freshwater resource utilization. However, the inefficiency of irrigation water usage has become a weak link in water resource management. To better assess irrigation water efficiency, an accurate estimation of regional irrigated areas is urgently needed. This study proposes a method for identifying actual irrigated areas based on remote sensing-derived evapotranspiration (ET) to address the challenge of accurately interpreting irrigated areas in tropical regions. Using Yunnan Province’s Yuanmou irrigation district as a case study, this research combined ground monitoring data and remote sensing data to identify actual irrigated areas through ET inversion and downscaling methods using the Penman–Monteith–Leuning (PML) model. In 2023, the total irrigated area interpreted from remote sensing in the study area was approximately 15,000 hm2, with a comparison against validation points revealing an extraction error of 16%. The small error indicates that this method can effectively enhance the reliability of monitoring actual irrigated areas, thus providing valuable data support for agricultural irrigation water management

STAT3-blocked whole-cell hepatoma vaccine induces cellular and humoral immune response against HCC

Abstract Background Whole-cell tumor vaccines have shown much promise; however, only limited success has been achieved for the goal of eliciting robust tumor-specific T-cell responses. Methods Hepatocellular carcinoma (HCC) cells, H22 and Hepa1–6, were modified by blocking the STAT3 signaling pathway with a STAT3 decoy oligodeoxynucleotide, and the immunogenicity and possibility of using these cell lysates as a vaccine were evaluated. Results STAT3-blocked whole HCC cell lysates inhibited tumor growth and tumorigenesis, and prolonged the survival of tumor-bearing mice. In addition, STAT3-blocked whole HCC cell lysates stimulated the activation of T cells and natural killer (NK) cells, and enhanced the infiltration of cytotoxic CD8+ T cells in the tumor tissues. In addition, the maturation of dendritic cells (DCs) was enhanced, which promoted the generation of immunological memory against HCC. Furthermore, secondary immune responses could be primed as soon as these immunized mice were challenged with HCC cells, accompanied by T cell and NK cell activation and infiltration. Additionally, immunization with this vaccine decreased the generation of Tregs and the production of TGF-β and IL-10. Importantly, STAT3-blocked whole HCC cell lysates prevented HCC-mediated exhaustion of T cells and NK cells, showing low expression of checkpoint molecules such as PD-1 and TIGIT on T cells and NK cells in the immunized mice. Conclusions The newly generated STAT3-blocked whole-cell HCC vaccine has potential for cancer cell vaccination

Involvement of Activation of PKR in HBx-siRNA-Mediated Innate Immune Effects on HBV Inhibition

RNA interference (RNAi) of virus-specific genes offers the possibility of developing a new anti-hepatitis B virus (anti-HBV) therapy. Recent studies have revealed that siRNAs can induce an innate immune response in vitro and in vivo. Here, HBVx (HBx) mRNA expression and HBV replication were significantly inhibited, followed by the enhancement of expression of type I interferons (IFNs), IFN-stimulated genes (ISG15 and ISG56) and proinflammatory cytokines after HepG2.2.15 cells were transfected with chemically synthesized HBx-siRNAs. Transfection with HBx-siRNAs also significantly increased expression of dsRNA-dependent protein kinase R (PKR) in HepG2.2.15 cells, followed by activation of downstream signaling events such as eukaryotic initiation factor 2a (eIF2-a). In PKR-over-expressing HepG2.2.15 cells, HBx-siRNAs exerted more potent inhibitory effects on HBV replication and greater production of type I IFNs. By contrast, the inhibitory effect of HBx-siRNAs on HBV replication was attenuated when PKR was inhibited or silenced, demonstrating that HBx-siRNAs greatly promoted PKR activation, leading to the higher production of type I IFN. Therefore, we concluded that PKR is involved in the innate immune effects mediated by HBx-siRNAs and further contributes to HBV inhibition. The bifunctional siRNAs with both gen

Estimation of Evapotranspiration Based on a Modified Penman–Monteith–Leuning Model Using Surface and Root Zone Soil Moisture

Most of the current parameterization schemes for the Penman–Monteith–Leuning evapotranspiration (ET) model (PML) consider meteorological and energy factors and land use types, but the analysis of the effect of soil moisture (SM) changes on ET processes lacks sufficient attention. This paper proposes a parameterization scheme for the sensitive parameters of the PML model considering soil water content, i.e., coupling the land surface SM in the calculation of soil evaporation coefficient f and coupling the SM of the root zone layer in the calculation of maximum stomatal conductance gsx, respectively. The new parameterization scheme is validated at 13 flux sites worldwide and showed significant improvements in improving the correlation with measured values. Moreover, based on the analysis of the spatial distribution of soil evaporation and vegetation transpiration, and the correlation between SM and ET, the regional characteristics of the effect of SM on ET are further revealed. This study provides a new idea for conducting the fusion simulation of SM based on a PML model, which is useful for the subsequent development of the model