Evidence of Urtica dioica Agglutinin's Antiproliferative and Anti-migratory Potentials on the Hyaluronic Acid-Overexpressing Prostate Cancer Cells.

Hyaluronic acid is composed of repeating sugar units, glucuronic acid and N-acetylglucosamine, which are often associated with increased tumor progression. Urtica dioica agglutinin is a potential component that exhibits a high affinity for binding to N-acetylglucosamine. This study aimed to investigate U. dioica Agglutinin's potential to inhibit the proliferation and migration of prostate cancer cells with high expression of hyaluronic acid through molecular docking and in vitro studies. The expression of hyaluronan synthase genes in prostate tissue and cell lines was checked by an in silico study, and the interaction between hyaluronic acid with both CD44 transmembrane glycoprotein and U. dioica agglutinin was analyzed through molecular docking. U. dioica Agglutinin's effect on cell viability (neutral red uptake assay), migration (scratch wound healing assays), and both CD44 and Nanog expression (quantitative real-time polymerase chain reaction) were assessed in vitro. The results showed that in prostate cancer cell lines, the PC3 cell line has the highest expression of hyaluronan synthase genes. U. dioica agglutinin exhibits an interaction of six specific residues on CD44 compared to hyaluronic acid's singular residue. While U. dioica agglutinin alone effectively reduced cell viability and wound closer (≥ 150 µg/mL), combining it with hyaluronic acid significantly shifted the effective concentration to a higher dose (≥ 350 µg/mL). These results, together with low Nanog and high CD44 gene expression, suggest that U. dioica agglutinin may impair the CD44-HA pathway in PC3 cells. This possibility is supported by U. dioica Agglutinin's ability to compete with hyaluronic acid for binding to CD44. Based on this, U. dioica agglutinin as a plant lectin shows promise in inhibiting cancer proliferation and migration by targeting its dependence on hyaluronic acid.

Missense polymorphisms potentially involved in mandibular prognathism.

Objective: The current study aimed to identify and analyze missense single nucleotide polymorphisms (SNPs) that can potentially cause mandibular prognathism. Methods: After reviewing the articles, 56 genes associated with mandibular prognathism were identified and their missense SNPs were retrieved from the NCBI website. Several web-based tools including CADD, PolyPhen-2, PROVEAN, SNAP2, PANTHER, FATHMM, and PON-P2 were used to filter out harmful SNPs. Additionally, ConSurf determined the level of evolutionary conservation at positions where SNPs occur. I-Mutant2 and MUpro predicted the effect of SNPs on protein stability. Furthermore, to investigate the structural and functional changes of proteins, HOPE and LOMETS tools were utilized. Results: Based on predictions in at least four web-based tools, the results indicated that PLXNA2-rs4844658, DUSP6-rs2279574, and FBN3-rs33967815 are harmful. These SNPs are located at positions with variable or average conservation and have the potential to reduce the stability of their respective proteins. Moreover, they may impair protein activity by causing structural and functional changes. Conclusions: In this study, we identified PLXNA2-rs4844658, DUSP6-rs2279574, and FBN3-rs33967815 as potential risk factors for mandibular prognathism using several web-based tools. According to the possible roles of PLXNA2, DUSP6, and FBN3 proteins in ossification pathways, we recommend that these SNPs be investigated further in experimental research. Through such studies, we hope to gain a better understanding of the molecular mechanisms involved in mandible formation.

Optimization of Affinity Chromatography Based on Sepharose 4B- chitin for Rapid Purification of Urtica dioica Agglutinin.

Background: Today, numerous antimicrobial and anticancer properties have been reported for plant lectins due to their ability to bind to carbohydrates. The Urtica dioica agglutinin (UDA lectin) is a monomeric, small, and low molecular weight glycoprotein. It has attracted the attention of many researchers for identification, treatment, and other clinical purposes. Objectives: The aim of this study is the optimization of the chitin affinity chromatography based on Sepharose 4B (CNBr-activated Sepharose 4B) for the rapid purification of UDA lectin from Urtica dioica rhizome. Materials and Methods: The chitin ligands were dissolved in 40% Trichloroacetic acid and attached to Sepharose 4B according to the Amersham-Biosciences instructions. The attachment of the ligand to the Sepharose 4B beads was investigated by Fourier transform infrared (FTIR) spectroscopy. An acidic crude extract of nettle rhizome passes from chromatographic columns in two sizes with dimensions: 24 x 0.51 cm and 8.44 x 0.86 cm. Quantity and quality of purified lectin were calculated by the Bradford microplate method: SDS-PAGE gel electrophoresis and human erythrocyte cell (RBC) hemagglutination, respectively. Results: The analysis of FTIR spectrograms showed that major changes were observed in the fingerprint regions. Besides, due to the dissolution of Sepharose 4B and chitin in the aqueous phase, this difference was not significant in the Imine and Nitrile regions. On the other hand, the comparative results of purification chromatograms showed that increasing the column length causes a smaller half-width and increases the length of the purified peak. Also, it leads to high-quality purified UDA lectin, with a molecular weight of almost 12.5 kDa in gel electrophoresis. Hemagglutination activity on trypsinized red blood cells was displayed, and agglutination of purified UDA lectin started at least at 300 µg.mL-1 concentration. Conclusion: According to our findings, we suggested that dissolving chitin in the polar solvent of Trichloroacetic acid, using Sepharose 4B as the beads of a matrix, and increasing the column length might lead to a decrease in the half-width of the peak. These can increase the purity and concentration of purified UDA lectin, and speed up the purification process. These findings could be used by researchers to accelerate the purification of UDA lectin in other studies, dealing with drug delivery systems, ELISA techniques, and cell growth.

CD44 rs13347C>T Variants in 3'UTR and Prostate Neoplasms: A Case-control Study and Bioinformatics Approach.

CD44, a cell-surface receptor and a key player in cellular signaling, can act as both tumor suppressor and promoter. This study aimed to investigate the association of CD44 rs13347C>T variants with prostate neoplasms, including both benign prostatic hyperplasia (BPH) and prostate cancers using a case-control and bioinformatics approach. Genomic DNA was extracted from 545 blood samples (225 BPH, 225 prostate cancers, and 95 control) and the CD44 rs13347C>T genotypes were identified using PCR-RFLP. We explored miRNA interactions using the miRNASNP-v3 database and GeneMANIA for co-expression networks. Results showed cancer patients had significantly higher PSA levels compared to both controls (p= 0.03) and BPH (p= 0.01). Additionally, digital rectal examination-positive and smoker BPH patients showed significantly the increased cancer risk (p= 0.004, p= 0.046). Prostate cancer group indicated significantly higher frequency of CD44 rs13347C>T mutant allele compared to control and BPH groups, particularly in TT and CT+TT genotypes (p < 0.05). miRNA SNP-v3 database predicted the mutant allele of CD44 rs13347C>T could lose 1 and gain 6 miRNAs for a new site created. Co-expression analysis revealed a direct interaction between CD44 and aryl hydrocarbon receptor (AHR), a gene known to be dysregulated in smokers. Furthermore, these genes alone display co-expression interactions with integrin subunit alpha 4 (ITGA4), protein plays a paradoxical role, both suppressing and promoting tumors. Based on the findings, the mutant allele of CD44 rs13347C>T may disrupt miRNA binding, which may potentially impact CD44, AHR, and ITGA4 expression in smokers, possibly contributing to prostate cancer progression.

Mutant Allele of CD44 (rs8193C>T) and Pum2 Regulatory Element as A Prognosis Factor of Prostate Neoplasms: A Case-Control and In Silico Studies.

OBJECTIVE: Expression of CD44 variant 6 (CD44v6) as a homing-associated cell adhesion molecule (HCAM), has proved to change most cancer cells. Aim of the study is the effect of mutant allele of CD44 (rs8193C>T) and Pum2 regulatory element as a prognosis factor of prostate neoplasms: a case-control and in silico studies in the Mazandaran province-Iran. MATERIALS AND METHODS: In a case-control study, CD44-rs8193C>T genotyping of the 420 prostate neoplasms (210 benign prostatic hyperplasia (BPH) patients and 210 prostate cancer patients) and 150 healthy samples are performed by the touchdown polymerase chain reaction with confronting two-pair primers (PCR-CTPP) method. The T mutant allele effects on the mRNA structure and cell pathways were also investigated in silico methods. RESULTS: Our results showed that the increase of T mutant allele frequency was significantly associated with BPH compared with prostate cancer. Furthermore, results showed TT genotype was significantly associated with BPH [odds ratio (OR)=0.572 and P=0.015], and also influenced the CD44v6 transcript secondary structure, miRNA binding, and regulatory element-binding site for Pum2 protein. Attachment of Pum2 to standard CD44 transcript may lead to transcript isoform-switching and shift-expression to a variety of CD44 isoforms, which can trigger some of the cell signaling pathways, such as Nanog-Stat, PKC-Nanog, and PKC-Twist. CONCLUSION: Based on this, the presence of the T mutant allele of CD44 (rs8193C>T) in the populations may create a regulatory element-binding site for Pum2. So, it could be known as a prognosis factor and prediction of prostate neoplasms. However, more comprehensive studies in different populations (with various ethnicities and large population sizes), and also CD44v6 gene expression studies in protein and transcript levels are required to confirm our data.

In Silico Analysis of Collagens Missense SNPs and Human Abnormalities.

Collagens are the most abundant proteins in the extra cellular matrix/ECM of human tissues that are encoded by different genes. There are single nucleotide polymorphisms/SNPs which are considered as the most useful biomarkers for some disease diagnosis or prognosis. The aim of this study is screening and identifying the functional missense SNPs of human ECM-collagens and investigating their correlation with human abnormalities. All of the missense SNPs were retrieved from the NCBI SNP database and screened for a global frequency of more than 0.1. Seventy missense SNPs that met the screening criteria were characterized for functional and stability impact using six and three protein analysis tools, respectively. Next, HOPE and geneMANIA analysis tools were used to show the effect of SNPs on three-dimensional structure (3D) and physical interaction of proteins. Results showed that 13 missense SNPs (rs2070739, rs28381984, rs13424243, rs1800517, rs73868680, rs12488457, rs1353613, rs59021909, rs9830253, rs2228547, rs3753841, rs2855430, and rs970547), which are in nine different collagen genes, affect the structure and function of different collagen proteins. Among these polymorphisms, COL4A3-rs13424243 and COL6A6-rs59021909 were predicted as the most effective ones. On the other hand, designed mutated and native 3D of rs13424243 variant illustrated that it can disturb the protein motifs. Also, geneMANIA predicted that COL4A3 and COL6A6 are interacting with some proteins including: DDR1, COL6A1, COL11A2 and so on. Based on our findings, ECM-collagens functional SNPs are important and may be considered as a risk factor or molecular marker for human disorders in the future studies.