Lamin B1 mRNA Research Articles

Knockdown of LMNB1 Inhibits the Proliferation of Lung Adenocarcinoma Cells by Inducing DNA Damage and Cell Senescence.

BackgroundLung cancer has considerably high mortality and morbidity rate. Lung adenocarcinoma (LUAD) tissues highly express lamin B1 (LMNB1), compared with normal tissues. In this study, we knocked down LMNB1 in LUAD cells A549 and NCI-1299 to explore the effect of its inhibition on the proliferation of cells and the potential mechanism.MethodsUsing bioinformatics methods, we analyzed the specificity of LMNB1 mRNA expression level in LUAD and its effect on prognosis from TCGA data. SiRNAs were used to knock down LMNB1 in the A549 cell line, and the knockdown effect was identified by western blotting and qRT-PCR. Through CCK8 cell proliferation assay, wound healing assay, TRAP, cloning formation Assay, DNase I-TUNEL assay, ATAC-seq, immunofluorescence, FISH, in vivo mouse xenograft studies, etc, we evaluated the influence and mechanism of LMNB1 on LUAD cell line proliferation in vitro and in vivo.ResultsAccording to bioinformatics analysis, LMNB1 is substantially abundant in LUAD tissues and is associated with tumor stage and patient survival (P < 0.05). After silencing LMNB1, the rate of cell growth, wound healing, the number of transwells, and the number of cell colonies all decreased significantly (P < 0.01). With the decreased LMNB1 expression, H3K9me3 protein expression decreases, chromosome accessibility increases, P53, P21, P16 and γ-H2AX protein expression increases, and the number of senescence staining positive cells increases. At the same time, in vivo mouse xenograft experiments showed that the tumor volume of the LMNB1-silenced group was significantly reduced, compared to that of the control group (P < 0.01), and the proliferation biomarker Ki-67 level (P < 0.01) was considerably reduced.ConclusionsOverexpression of LMNB1 in LUAD cells is significant, which has excellent potential to be an indicator for evaluating the clinical prognosis of LUAD patients and a target for precise treatment.

Direct interaction analysis of microRNA-351-5p and nuclear scaffold lamin B1 mRNA by the cell-free in vitro mRNA/miRNA binding evaluation system

We previously demonstrated that miR-351-5p regulates nuclear scaffold lamin B1 expression and mediates the anticancer floxuridine-induced necrosis shift to apoptosis in mammalian tumor cells. Notably, it is unknown whether lamin B1 mRNA is a direct target of miR-351-5p. Here, we show that miR-351-5p interacts with a lamin B1 mRNA partial sequence by using the cell-free in vitro miRNA and mRNA binding evaluation system. In addition, the interaction of miR-351-5p/lamin B1 mRNA was suppressed by an miR-351-5p inhibitor. Our findings are important in exploring the functions of miRNAs in cellular processes, including cell death.

Tissue specificity of senescent cell accumulation during physiologic and accelerated aging of mice.

Senescent cells accumulate with age in vertebrates and promote aging largely through their senescence‐associated secretory phenotype (SASP). Many types of stress induce senescence, including genotoxic stress. ERCC1‐XPF is a DNA repair endonuclease required for multiple DNA repair mechanisms that protect the nuclear genome. Humans or mice with reduced expression of this enzyme age rapidly due to increased levels of spontaneous, genotoxic stress. Here, we asked whether this corresponds to an increased level of senescent cells. p16Ink4a and p21Cip1 mRNA were increased ~15‐fold in peripheral lymphocytes from 4‐ to 5‐month‐old Ercc1 −/∆ and 2.5‐year‐old wild‐type (WT) mice, suggesting that these animals exhibit a similar biological age. p16Ink4a and p21Cip1 mRNA were elevated in 10 of 13 tissues analyzed from 4‐ to 5‐month‐old Ercc1 −/∆ mice, indicating where endogenous DNA damage drives senescence in vivo. Aged WT mice had similar increases of p16Ink4a and p21Cip1 mRNA in the same 10 tissues as the mutant mice. Senescence‐associated β–galactosidase activity and p21Cip1 protein also were increased in tissues of the progeroid and aged mice, while Lamin B1 mRNA and protein levels were diminished. In Ercc1 −/Δ mice with a p16Ink4a luciferase reporter, bioluminescence rose steadily with age, particularly in lung, thymus, and pancreas. These data illustrate where senescence occurs with natural and accelerated aging in mice and the relative extent of senescence among tissues. Interestingly, senescence was greater in male mice until the end of life. The similarities between Ercc1 −/∆ and aged WT mice support the conclusion that the DNA repair‐deficient mice accurately model the age‐related accumulation of senescent cells, albeit six‐times faster.

Allele-specific silencing as treatment for gene duplication disorders: proof-of-principle in autosomal dominant leukodystrophy.

Allele-specific silencing by RNA interference (ASP-siRNA) holds promise as a therapeutic strategy for downregulating a single mutant allele with minimal suppression of the corresponding wild-type allele. This approach has been effectively used to target autosomal dominant mutations and single nucleotide polymorphisms linked with aberrantly expanded trinucleotide repeats. Here, we propose ASP-siRNA as a preferable choice to target duplicated disease genes, avoiding potentially harmful excessive downregulation. As a proof-of-concept, we studied autosomal dominant adult-onset demyelinating leukodystrophy (ADLD) due to lamin B1 (LMNB1) duplication, a hereditary, progressive and fatal disorder affecting myelin in the CNS. Using a reporter system, we screened the most efficient ASP-siRNAs preferentially targeting one of the alleles at rs1051644 (average minor allele frequency: 0.45) located in the 3' untranslated region of the gene. We identified four siRNAs with a high efficacy and allele-specificity, which were tested in ADLD patient-derived fibroblasts. Three of the small interfering RNAs were highly selective for the target allele and restored both LMNB1 mRNA and protein levels close to control levels. Furthermore, small interfering RNA treatment abrogates the ADLD-specific phenotypes in fibroblasts and in two disease-relevant cellular models: murine oligodendrocytes overexpressing human LMNB1, and neurons directly reprogrammed from patients' fibroblasts. In conclusion, we demonstrated that ASP-silencing by RNA interference is a suitable and promising therapeutic option for ADLD. Moreover, our results have a broad translational value extending to several pathological conditions linked to gene-gain in copy number variations.

Differential Expression of LaminB1 in the Developing Rat Cochlea.

To explore the temporal expression pattern of LaminB1 in the cochlea of postnatal rat, and whether LaminB1 is associated with cochlear development. Sprague-Dawley rats ranging from postnatal day 0 (p0) to 21 (p21) were used. The tissues of stria vascularis (STV) including spiral ligament, spiral ganglion cell (SGC), and basilar membrane (BM), including the organ of Corti, were dissected, respectively. Immunofluorescence, quantitative real-time polymerase chain reaction, and western blot were applied to detect the expression of LaminB1 in individual cochlear tissues at both mRNA and protein levels. Immunofluorescence revealed that LaminB1 was localized in the outer hair cells, inner hair cells, Kolliker's organ, Reissner's membrane, SGC, STV, and spiral ligament. The intensity of staining surrounding the scala media decreased during cochlear development. The expression of LaminB1 mRNA and protein in STV, SGC, and BM was at a maximum level at p0 but gradually declined to a minimum level at p21. Our research provided direct evidence that LaminB1 was expressed in the developing cochlea and developmentally regulated in cochlear tissues, suggesting a possible role of LaminB1 in cochlear development. Our result provided a theoretical basis for further study about the physiological function of LaminB1 in the peripheral auditory system.

Analysis of Lamin B1, Vimentin and Anti-Ku86 as Prospective Biomarkers of Hepatocellular Carcinoma in Patients with Hepatitis C Virus Infection.

Hepatocellular carcinoma (HCC) is the fifth most common malignancy and most frequently develops in patients with cirrhosis. Surveillance strategies are recommended in high-risk groups because early detection of small lesions improves the likelihood of curative treatment. This study investigated the prospective clinical significance of serum levels of anti-Ku86 and plasma levels of lamin B1and vimentin as early markers of HCC. We recruited 74 patients at Assiut University Hospital-37 with HCC and 37 with chronic liver disease (liver cirrhosis patients)-and 36 age- and sex-matched healthy controls. Lamin B1 and vimentin mRNA expression levels were evaluated by reverse transcription-PCR and serum levels of anti-Ku86 were measured by enzyme-linked immunosorbent assay. Compared with liver disease patients and controls, HCC patients showed higher levels of lamin B1 mRNA (sensitivity, 96%; specificity, 65%), vimentin mRNA (sensitivity, 94%; specificity, 92%), and anti-Ku86 (sensitivity, 94%; specificity, 80%). LaminB1 levels were significantly higher in patients with a tumor size < 2 cm than in patients with tumors 2-5 cm and >5cm in size. Lamin B1 had significant positive correlations with alpha-fetoprotein (AFP) (P=0.034) and anti-Ku86 (P=0.002). Receiver operating characteristic curves for differentiating HCCfrom liver cirrhosis revealed a higher area under the curve(AUC).for vimentin than for AFP, lamin B1, and anti-Ku86 for the diagnosis of HCC (P<0.001). Circulating levels of anti-Ku86, lamin B1,and vimentin might be potential surrogate markers of HCC, either alone or in combination with AFP. However, independent and discriminative serological biomarkers with higher sensitivity and specificity are still needed for the early detection of HCC.

Using Lamin B1 mRNA for the early diagnosis of hepatocellular carcinoma: a cross-sectional diagnostic accuracy study

Background : Hepatocellular carcinoma (HCC) is vital medical issue in Egypt. It accounts for 70.48% of all liver tumors among Egyptians. The aim of this study was to determine the diagnostic role of plasma levels of mRNA of lamin B1 by RT-qPCR as an early marker of HCC. Methods: This study was conducted at the Clinical Pathology Department in collaboration with the Department of Tropical Medicine and Infectious Diseases at Ain Shams University Hospitals. It included 30 patients with primary HCC and viral cirrhosis (all were hepatitis C virus-positive) (Group I), in addition to 10 patients with chronic liver diseases (Group II) and 10 healthy age- and sex-matched subjects (Group III). Group I was further classified according to the Barcelona-Clinic Liver Cancer Staging System. Serum α-fetoprotein (AFP) chemiluminescent-immunoassays and RT-qPCR analysis of plasma lamin B1 mRNA levels were performed for all participants. Results: AFP and lamin B1 significantly elevated in patients with HCC compared to those in the other studied groups. AFP and lamin B1 status could discriminate group I from group II and III. A significant increase was found among the three Barcelona stages with regards to AFP and lamin B1 levels. A significant decrease was found between group II and stage 0, A and B with regards to AFP and lamin B1. Lamin B1 and AFP could both differentiate HCC patients with one tumor nodule (T1) from those with two or more tumor nodules (T2&Tm), as well as between those with tumor sizes >3 cm and ≤3 cm. Conclusion: Measurement of lamin B1 mRNA is recommended in patients with chronic liver disease with normal serum AFP, especially in known cirrhotic patients that deteriorate rapidly without any apparent etiology.

Using Lamin B1 mRNA for the early diagnosis of hepatocellular carcinoma: a cross-sectional diagnostic accuracy study.

Background: Hepatocellular carcinoma (HCC) is vital medical issue in Egypt. It accounts for 70.48% of all liver tumors among Egyptians. The aim of this study was to determine the diagnostic role of plasma levels of mRNA of lamin B1 by RT-qPCR as an early marker of HCC. Methods: This study was conducted at the Clinical Pathology Department in collaboration with the Department of Tropical Medicine and Infectious Diseases at Ain Shams University Hospitals. It included 30 patients with primary HCC and viral cirrhosis (all were hepatitis C virus-positive) (Group I), in addition to 10 patients with chronic liver diseases (Group II) and 10 healthy age- and sex-matched subjects (Group III). Group I was further classified according to the Barcelona-Clinic Liver Cancer Staging System. Serum α-fetoprotein (AFP) chemiluminescent-immunoassays and RT-qPCR analysis of plasma lamin B1 mRNA levels were performed for all participants. Results: AFP and lamin B1 significantly elevated in patients with HCC compared to those in the other studied groups. AFP and lamin B1 status could discriminate group I from group II and III. A significant increase was found among the three Barcelona stages with regards to AFP and lamin B1 levels. A significant decrease was found between group II and stage 0, A and B with regards to AFP and lamin B1. Lamin B1 and AFP could both differentiate HCC patients with one tumor nodule (T1) from those with two or more tumor nodules (T2&Tm), as well as between those with tumor sizes >3 cm and ≤3 cm. Conclusion: Measurement of lamin B1 mRNA is recommended in patients with chronic liver disease with normal serum AFP, especially in known cirrhotic patients that deteriorate rapidly without any apparent etiology.

Protective Effect of Copper (I)-Nicotinate Complex on DAB-Induced Hepatocellular Carcinoma in Rats

The aim of the present work was to evaluate the effect of copper (I)-nicotinate complex (CNC) on experimentally 4-Dimethylaminoazobenzene (DAB)-induced hepatocellular carcinoma (HCC) in male rats. In addition to the histopathological examination we measured hepatic glutathione content, malondialdehyde, nitric oxide levels, lamin B1 m-RNA, caspase-3 activity and serum interleukin-12 level after 2, 4, 6, 8 months from commencement of the experiment. Results: histopathological examination showed HCC development after 4 months of DAB administration. The hepatic glutathione content, lamin B1 m-RNA and nitric oxide levels were significantly elevated, while malondialdehyde, interlukin-12 levels and caspase-3 activity were significantly decreased with the progression and development of the HCC. On the other hand, administration of CNC one month before DAB delayed the development of neoplastic growth to the 8th month. Interestingly, when CNC was administered one month after DAB, it successfully prevented HCC development throughout the whole experiment as confirmed by histopathological data and explained by biochemical markers, as glutathione, lamin B-1 and nitric oxide were significantly declined but malondialdehyde, interlukin-12 and caspase-3 activity were significantly elevated compared to that in corresponding control group. Conclusion: CNC was able to delay or prevent HCC development in rats fed with the potent liver carcinogen DAB. Our data shows that CNC exerts its anti-tumour effects through modulating oxidative stress status as well as the machinery of apoptosis and angiogenesis. Therefore, CNC may be used as a potential protective anticancer agent.

Decreased Dp71 expression is associated with gastric adenocarcinoma prognosis.

For the first time, dramatically decreased Dp71 protein and mRNA was found in 34 pairs of resected primary gastric adenocarcinoma. Immunohistochemistry identified Dp71 expression suppressed in 72.2% of 104 gastric cancer patients. The decreased Dp71 expression was significantly correlated with cancer differentiation (P=0.001) and lymph vascular invasion (p=0.041). Decreased Dp71 expression was associated with a poor gastric adenocarcinoma prognosis (P=0.001). Significantly less Dp71 mRNA and protein were found in BGC823, SGC7901, AGS compared with GES-1. Via increasing lamin B1 mRNA and protein, enforced Dp71d and Dp71f expression resulted in SGC7901 proliferation inhibition. Co-IP proved interaction of Dp71 with lamin B1 in GES-1 cells. Further expression characterization showed reduced lamin B1 in gastric cancer tissue and cancer cells. Increasing lamin B1 expression results in the growth inhibition of SGC7901, which suggests that Dp71-lamin B1 protein complex plays an important role for the newly identified tumor suppressive function of Dp71.

Detection of circulating lamin B1 and glypican-3 for early diagnosis of hepatocellular carcinoma patients

Objective: To assess circulating lamin B1 (LMNB1) mRNA and glypican-3 cytotoxic T lymphocytes (CTLs) as markers for early detection of hepatocellular carcinoma (HCC). Background: HCC is the fifth most common form of cancer worldwide and the third most common cause of cancer-related deaths. HCC often occurs in the background of a cirrhotic liver. In recent years, surveillance strategies in patients at a higher risk of HCC have led to the diagnosis of the disease at much earlier stages. Patients in early stages have a much higher chance of curative response with different treatment options. Materials and methods: The study included 50 participates, of which 13 were patients with posthepatitis C liver cirrhosis (group I), 13 were patients with early stages of HCC (group IIa), 14 were patients with late stages of HCC (group IIb), and 10 were healthy volunteers with matched age and sex as a control group (group III). The patients were selected from the outpatient clinic and inpatient department of the National Liver Institute, Menoufiya University. LMNB1 mRNA was assessed by reverse transcription-PCR and glypican3 CTLs by flow cytometry and then we compared that with α±-feto protein. Results: Glypican3 CTLs can be used for early detection of HCC patients with 100% specificity and sensitivity. In addition, LMNB1 mRNA can be used in early detection of HCC with specificity and sensitivity 92 and 100%, respectively. Conclusion: Circulating LMNB1 mRNA and glypican3 CTLs can be used as markers for early detection of HCC patients.

Circulating mRNA Lamin B1 for Detection of Early Stages of Hepatocellular Carcinoma

Background: Hepatocel lular carcinoma (HCC) is the fi fth most common cancer and the thi rd leading cause ofdeath worldwide, causing 600,000 deaths annual ly . In Egypt, HCC accounts for 4.7% of al l l iver diseases.Hospi tal -based studies have reported an overal l increase in the relative frequency of al l l iver cancers in Egyptfrom approximately 4% in 1993 to 7.3% in 2003, more than 95% of which being HCC . Patients diagnosed at anearly stage of HCC have the best prognosis, where resection and t ransplantation achieve the best outcomes inwel l -selected candidates, wi th a 5-year - survival rate of 50 to 70%. In this regard, the present study aimed todemonstrate the potential cl inical importance of plasma levels of ci rculating mRNA of lamin B1 by real -time RTPCR as an early marker of HCC. .Methods: This study was conducted at the Cl inical Pathology Department and the subjects were selected fromDepartment of Tropical Medicine and Infectious Diseases at Ain Shams Universi ty Hospi tals. It included thi r ty (30)HCC patients of di fferent stages (group I) , they were further divided into 3 stages using BCLC staging system intostage A, B and C, in addi tion to ten (10) cases wi th benign l iver diseases serving as patient cont rols (group II ) andten (10) heal thy age-and sex -matched subjects serving as heal thy cont rols (group III ) .Results: The plasma laminB1 mRNA showed a much bet ter performance to di fferentiate between HCC cases andthe 2 control groups, where i ts sensi tivi ty was 100% and speci fici ty was 90%.Conclusions: Addi tion of plasma lamin B1 mRNA to the cur rent standard tests for diagnosis of HCC as a newdiagnostic and screening tool . This, in turn, could greatly improve the abi l i ty to identi fy such patients and thuscould al low them to benefi t f rom earl ier t reatme nt.

The clinicopathological significance of lamin A/C, lamin B1 and lamin B receptor mRNA expression in human breast cancer

Lamin A/C (LMNA), lamin B1 (LMNB1) and lamin B receptor (LBR) have key roles in nuclear structural integrity and chromosomal stability. In this study, we have studied the relationships between the mRNA expressions of A-type lamins, LMNB1 and LBR and the clinicopathological parameters in human breast cancer. Samples of breast cancer tissues (n = 115) and associated non-cancerous tissue (ANCT; n = 30) were assessed using reverse transcription and quantitative PCR. Transcript levels were correlated with clinicopathological data. Higher levels of A-type lamins and LMNB1 mRNA expression were seen in ANCT. Higher lamin A/C expression was associated with the early clinical stage (TNM1 vs. TNM3 — 13 vs. 0.21; p = 0.0515), with better clinical outcomes (disease-free survival vs. mortality — 11 vs. 1; p = 0.0326), and with better overall (p = 0.004) and disease-free survival (p = 0.062). The expression of LMNB1 declined with worsening clinical outcome (disease-free vs. mortalities — 0.0011 vs. 0.000; p = 0.0177). LBR mRNA expression was directly associated with tumor grade (grade 1 vs. grade 3 — 0.00 vs. 0.00; p = 0.0479) and Nottingham Prognostic Index (NPI1 vs. NPI3 — 0.00 vs. 0.00; p = 0.0551). To the best of our knowledge, this is the first study to suggest such a role for A-type lamins, lamin B1 and LBR in human breast cancer, identifying an important area for further research.

Translational control of nuclear lamin B1 mRNA during oogenesis and early development of Xenopus

Cytoplasmic polyadenylation of specific mRNAs is commonly correlated with their translational activation during development. A canonical nuclear polyadenylation element AAUAAA (NPE) and cytoplasmic polyadenylation element(s) (CPE) are necessary and sufficient for polyadenylation during egg maturation. We have characterized cis-acting sequences of Xenopus nuclear lamin B1 mRNA that mediate translational regulation. By injection of synthetic RNAs into oocytes we show that the two CPE-like elements found in the 3′-untranslated region of B1 mRNA act as translational repressors in oocytes. The same CPEs in conjunction with the NPE confer transient polyadenylation and translational activation during egg maturation. Poly(A) length determination of the endogenous lamin B1 mRNA reveals a gradual increase of poly(A) tail length in early development up to mid-blastula, and a shortening of poly(A) tails during gastrulation and neurulation. The same kinetic and extent of polyadenylation and poly(A) tail shortening is observed with synthetic RNAs injected into fertilized eggs. Polyadenylation and translational activation of these RNAs is independent of the two CPEs and a NPE during early development. While translational regulation of lamin B1 mRNA functions in parts via established mechanisms, the pattern of polyadenylation and deadenylation during early development points to a novel mode of translational regulation.