Large-volume fully automated cell reconstruction generates a cell atlas of plant tissues.

The geometric shape and arrangement of individual cells play a role in shaping organ functions. However, analyzing multicellular features and exploring their connectomes in centimeter-scale plant organs remain challenging. Here, we established a set of frameworks named Large-Volume Fully Automated Cell Reconstruction (LVACR), enabling the exploration of three-dimensional (3D) cytological features and cellular connectivity in plant tissues. Through benchmark testing, our framework demonstrated superior efficiency in cell segmentation and aggregation, successfully addressing the inherent challenges posed by light sheet fluorescence microscopy (LSFM) imaging. Using LVACR, we successfully established a cell atlas of different plant tissues. Cellular morphology analysis revealed differences of cell clusters and shapes in between different poplar (P. simonii Carr. and P. canadensis Moench.) seeds, whereas topological analysis revealed that they maintained conserved cellular connectivity. Furthermore, LVACR spatiotemporally demonstrated an initial burst of cell proliferation, accompanied by morphological transformations at an early stage in developing the shoot apical meristem. During subsequent development, cell differentiation produced anisotropic features, thereby resulting in various cell shapes. Overall, our findings provided valuable insights into the precise spatial arrangement and cellular behavior of multicellular organisms, thus enhancing our understanding of the complex processes underlying plant growth and differentiation.

Three-dimensional reconstruction and multiomics analysis reveal a unique pattern of embryogenesis in Ginkgo biloba.

Ginkgo (Ginkgo biloba L.) is one of the earliest extant species in seed plant phylogeny. Embryo development patterns can provide fundamental evidence for the origin, evolution, and adaptation of seeds. However, the architectural and morphological dynamics during embryogenesis in G. biloba remain elusive. Herein, we obtained over 2,200 visual slices from 3 stages of embryo development using micro-computed tomography imaging with improved staining methods. Based on 3-dimensional (3D) spatiotemporal pattern analysis, we found that a shoot apical meristem with 7 highly differentiated leaf primordia, including apical and axillary leaf buds, is present in mature Ginkgo embryos. 3D rendering from the front, top, and side views showed 2 separate transport systems of tracheids located in the hypocotyl and cotyledon, representing a unique pattern of embryogenesis. Furthermore, the morphological dynamic analysis of secretory cavities indicated their strong association with cotyledons during development. In addition, we identified genes GbLBD25a (lateral organ boundaries domain 25a), GbCESA2a (cellulose synthase 2a), GbMYB74c (myeloblastosis 74c), GbPIN2 (PIN-FORMED 2) associated with vascular development regulation, and GbWRKY1 (WRKYGOK 1), GbbHLH12a (basic helix-loop-helix 12a), and GbJAZ4 (jasmonate zim-domain 4) potentially involved in the formation of secretory cavities. Moreover, we found that flavonoid accumulation in mature embryos could enhance postgerminative growth and seedling establishment in harsh environments. Our 3D spatial reconstruction technique combined with multiomics analysis opens avenues for investigating developmental architecture and molecular mechanisms during embryogenesis and lays the foundation for evolutionary studies of embryo development and maturation.

The G allele of SNP rs3922 reduces the binding affinity between IGF2BP3 and CXCR5 correlating with a lower antibody production.

Effective vaccines that function through humoral immunity seek to produce high-affinity antibodies. Our previous research identified the single-nucleotide polymorphism rs3922G in the 3'UTR of CXCR5 as being associated with nonresponsiveness to the hepatitis B vaccine. The differential expression of CXCR5 between the dark zone (DZ) and light zone (LZ) is critical for organizing the functional structure of the germinal center (GC). In this study, we report that the RNA-binding protein IGF2BP3 can bind to CXCR5 mRNA containing the rs3922 variant to promote its degradation via the nonsense-mediated mRNA decay pathway. Deficiency of IGF2BP3 leads to increased CXCR5 expression, which results in the disappearance of CXCR5 differential expression between DZ and LZ, disorganized GCs, aberrant somatic hypermutations, and reduced production of high-affinity antibodies. Furthermore, the affinity of IGF2BP3 for the rs3922G-containing sequence is lower than that for the rs3922A counterpart, which may explain the nonresponsiveness to the hepatitis B vaccination. Together, our findings suggest that IGF2BP3 plays a crucial role in the production of high-affinity antibodies in the GC by binding to the rs3922-containing sequence to regulate CXCR5 expression.

Analyses of high spatial resolution datasets identify genes associated with multi-layered secondary cell wall thickening in Pinus bungeana.

BACKGROUND AND AIMS: Secondary cell wall (SCW) thickening is a major cellular developmental stage determining wood structure and properties. Although the molecular regulation of cell wall deposition during tracheary element differentiation has been well established in primary growth systems, less is known about the gene regulatory processes involved in the multi-layered SCW thickening of mature trees. METHODS: Using third-generation [long-read single-molecule real-time (SMRT)] and second-generation [short-read sequencing by synthesis (SBS)] sequencing methods, we established a Pinus bungeana transcriptome resource with comprehensive functional and structural annotation for the first time. Using these approaches, we generated high spatial resolution datasets for the vascular cambium, xylem expansion regions, early SCW thickening, late SCW thickening and mature xylem tissues of 71-year-old Pinus bungeana trees. KEY RESULTS: A total of 79 390 non-redundant transcripts, 31 808 long non-coding RNAs and 5147 transcription factors were annotated and quantified in different xylem tissues at all growth and differentiation stages. Furthermore, using this high spatial resolution dataset, we established a comprehensive transcriptomic profile and found that members of the NAC, WRKY, SUS, CESA and LAC gene families are major players in early SCW formation in tracheids, whereas members of the MYB and LBD transcription factor families are highly expressed during late SCW thickening. CONCLUSIONS: Our results provide new molecular insights into the regulation of multi-layered SCW thickening in conifers. The high spatial resolution datasets provided can serve as important gene resources for improving softwoods.

The Ring-Closing Reaction of Cyclopentadiene Probed with Ultrafast X-ray Scattering.

The dynamics of cyclopentadiene (CP) following optical excitation at 243 nm was investigated by time-resolved pump-probe X-ray scattering using 16.2 keV X-rays at the Linac Coherent Light Source (LCLS). We present the first ultrafast structural evidence that the reaction leads directly to the formation of bicyclo[2.1.0]pentene (BP), a strained molecule with three- and four-membered rings. The bicyclic compound decays via a thermal backreaction to the vibrationally hot CP with a time constant of 21 ± 3 ps. A minor channel leads to ring-opened structures on a subpicosecond time scale.

Safety, pharmacokinetics, and pharmacodynamics of SHR7280, an oral gonadotropin-releasing hormone receptor antagonist, in healthy men: a randomized, double-blind, placebo-controlled phase 1 study.

BACKGROUND: Gonadotropin-releasing hormone (GnRH) antagonists are a promising therapeutic approach for treating hormone-dependent prostate cancer. Currently, the mainstream GnRH antagonists are polypeptide agents administered through subcutaneous injection. In this study, we evaluated the safety, pharmacokinetics (PK), and pharmacodynamics (PD) of SHR7280, an oral small molecule GnRH antagonist, in healthy men. METHODS: This phase 1 trial was a randomized, double-blind, placebo-controlled, and dose-ascending study. Eligible healthy men were randomized in a 4:1 ratio to receive either oral SHR7280 tablets or placebo twice daily (BID) for 14 consecutive days. The SHR7280 dose was initiated at 100 mg BID and then sequentially increased to 200, 350, 500, 600, 800, and 1000 mg BID. Safety, PK, and PD parameters were assessed. RESULTS: A total of 70 subjects were enrolled and received the assigned drug, including 56 with SHR7280 and 14 with placebo. SHR7280 was well-tolerated. The incidence of adverse events (AEs, 76.8% vs 85.7%) and treatment-related AEs (75.0% vs 85.7%), as well as the severity of AEs (moderate AEs, 1.8% vs 7.1%) were similar between the SHR7280 group and placebo group. SHR7280 was rapidly absorbed in a dose-dependent manner, with a median Tmax of each dose group ranging from 0.8 to 1.0 h on day 14 and a mean t1/2 ranging from 2.8 to 3.4 h. The PD results demonstrated that SHR7280 exhibited a rapid and dose-proportional suppression of hormones, including LH, FSH, and testosterone, with maximum suppression achieved at doses of 800 and 1000 mg BID. CONCLUSIONS: SHR7280 showed an acceptable safety profile, as well as favorable PK and PD profiles within a dose range of 100 to 1000 mg BID. This study proposes a rationale for further investigation of SHR7280 as a potential androgen deprivation therapy. TRIAL REGISTRATION: Clinical trials.gov NCT04554043; registered September 18, 2020.

Spatial organization and connectivity of wood rays in Pinus massoniana xylem based on high-resolution µCT-assisted network analysis.

MAIN CONCLUSION: Spatial organization and connectivity of wood rays in Pinus massoniana was comprehensively viewed and regarded as anatomical adaptions to ensure the properties of rays in xylem. Spatial organization and connectivity of wood rays are essential for understanding the wood hierarchical architecture, but the spatial information is ambiguous due to small cell size. Herein, 3D visualization of rays in Pinus massoniana was performed using high-resolution µCT. We found brick-shaped rays were 6.5% in volume fractions, nearly twice the area fractions estimated by 2D levels. Uniseriate rays became taller and wider during the transition from earlywood to latewood, which was mainly contributed from the height increment of ray tracheids and widened ray parenchyma cells. Furthermore, both volume and surface area of ray parenchyma cells were larger than ray tracheids, so ray parenchyma took a higher proportion in rays. Moreover, three different types of pits for connectivity were segmented and revealed. Pits in both axial tracheids and ray tracheids were bordered, but the pit volume and pit aperture of earlywood axial tracheids were almost tenfold and over fourfold larger than ray tracheids. Contrarily, cross-field pits between ray parenchyma and axial tracheids were window-like with the principal axis of 31.0 µm, but its pit volume was approximately one-third of axial tracheids. Additionally, spatial organization of rays and axial resin canal was analyzed by a curved surface reformation tool, providing the first evidence of rays close to epithelial cells inward through the resin canal. Epithelial cells had various morphologies and large variations in cell size. Our results give new insights into the organization of radial system of xylem, especially the connectivity of rays with adjacent cells.

Analysis of the clinical features and risk factors of device-related pressure injuries in the operating room.

To describe the clinical features and risk factors of device-related pressure injuries (DRPIs) in the operating room. The clinical features of the DRPIs in patients undergoing elective surgery in a tertiary hospital in 2020 were investigated through prospective data collection. A DRPI-related questionnaire was designed for the patients, and those who did not experience any DRPI were selected according to a ratio of 1:2. Logistic regression analysis was performed in terms of the independent risk factors of operating-room DRPIs. A P-value of <.05 indicated a statistically significant difference. The incidence of operating-room DRPIs was 0.56%, and the proportion of stage I injuries was 73.53%. The injury-related devices included vital monitoring devices (31.62%), auxiliary therapy devices (27.94%), therapy devices (19.12%), and dressings (3.67%). Non-bone protuberances, such as the upper arms and thighs, were common injury sites. The patients' body mass index, mean arterial pressure, and instrument action time were independent risk factors for the operating-room DRPIs. To reduce the incidence of operating-room DRPIs, it is of great clinical significance to focus on the characteristics of the surgical patients and the types of surgery-related devices used and to take personalised preventive measures based on the relevant risk factors.

Associations between the TNMD rs4828038 and ACE2 rs879922 polymorphisms and preeclampsia susceptibility: a case-control study.

A case-control study was designed to investigate the association between the angiotensin converting enzyme 2 (ACE2) rs879922, glucose-6-phosphate dehydrogenase (G6PD) rs1050828, and tenomodulin (TNMD) rs4828038 single nucleotide polymorphisms (SNPs), and preeclampsia. A total of 356 Han Chinese pregnant women (170 controls and 186 cases) were recruited into the study. ACE2 rs879922, G6PD rs1050828, and TNMD rs4828038 were tested by the targeted next-generation sequencing technology and the data were analyzed using SPSS version 18. Genotyping of results revealed that patients with the CC/CT genotype in SNP rs4828038 or CC/CG genotype in SNP rs879922 had a significantly decreased susceptibility to late-onset preeclampsia (CC/CT versus TT: OR = 0.543, 95% CI = 0.378 to 0.779, p = .001; CC/CG versus GG: OR = 0.510, 95% CI = 0.038 to 0.860, p = .012). Our study found that the polymorphisms TNMD rs4828038 and ACE2 rs879922 might be associated with late-onset preeclampsia.IMPACT STATEMENTWhat is already known on this subject? Preeclampsia is associated with multiple SNPs, and ACE2 rs879922, G6PD rs1050828, and TNMD rs4828038 are related to essential hypertension and glucose and lipid metabolism disorders. Essential hypertension, diabetes, and dyslipidemia are risks for preeclampsia. The associations between those three SNPs and preeclampsia have not been reported.What do the results of this study add? The polymorphisms of TNMD rs4828038 and ACE2 rs879922 might be associated with the risk of late-onset preeclampsia. There was no relationship between SNP rs1050828 and preeclampsia.What are the implications of these findings for clinical practice and/or further research? TNMD rs4828038 and ACE2 rs879922 might be target sites for genetic diagnosis and therapy, and the levels of mRNA and protein in pregnant women with preeclampsia should be further tested.

Mapping static core-holes and ring-currents with X-ray scattering.

Measuring the attosecond movement of electrons in molecules is challenging due to the high temporal and spatial resolutions required. X-ray scattering-based methods are promising, but many questions remain concerning the sensitivity of the scattering signals to changes in density, as well as the means of reconstructing the dynamics from these signals. In this paper, we present simulations of stationary core-holes and electron dynamics following inner-shell ionization of the oxazole molecule. Using a combination of time-dependent density functional theory simulations along with X-ray scattering theory, we demonstrate that the sudden core-hole ionization produces a significant change in the X-ray scattering response and how the electron currents across the molecule should manifest as measurable modulations to the time dependent X-ray scattering signal. This suggests that X-ray scattering is a viable probe for measuring electronic processes at time scales faster than nuclear motion.

Determination of excited state molecular structures from time-resolved gas-phase X-ray scattering.

We present a comprehensive investigation of a recently introduced method to determine transient structures of molecules in excited electronic states with sub-ångstrom resolution from time-resolved gas-phase scattering signals. The method, which is examined using time-resolved X-ray scattering data measured on the molecule N-methylmorpholine (NMM) at the Linac Coherent Light Source (LCLS), compares the experimentally measured scattering patterns against the simulated patterns corresponding to a large pool of molecular structures to determine the full set of structural parameters. In addition, we examine the influence of vibrational state distributions and find the effect negligible within the current experimental detection limits, despite that the molecules have a comparatively high internal vibrational energy. The excited state structures determined using three structure pools generated using three different computational methods are in good agreement, demonstrating that the procedure is largely independent of the computational chemistry method employed as long as the pool is sufficiently expansive in the vicinity of the sought structure and dense enough to yield good matches to the experimental patterns.

Strong-field induced fragmentation and isomerization of toluene probed by ultrafast femtosecond electron diffraction and mass spectrometry.

We investigate the fragmentation and isomerization of toluene molecules induced by strong-field ionization with a femtosecond near-infrared laser pulse. Momentum-resolved coincidence time-of-flight ion mass spectrometry is used to determine the relative yield of different ionic products and fragmentation channels as a function of laser intensity. Ultrafast electron diffraction is used to capture the structure of the ions formed on a picosecond time scale by comparing the diffraction signal with theoretical predictions. Through the combination of the two measurements and theory, we are able to determine the main fragmentation channels and to distinguish between ions with identical mass but different structures. In addition, our diffraction measurements show that the independent atom model, which is widely used to analyze electron diffraction patterns, is not a good approximation for diffraction from ions. We show that the diffraction data is in very good agreement with ab initio scattering calculations.

miR-34a regulates lipid metabolism by targeting SIRT1 in non-alcoholic fatty liver disease with iron overload.

BACKGROUND: Micro-ribonucleic acids (miRNAs) have been implicated in the regulation of non-alcoholic fatty liver disease (NAFLD), a leading cause of chronic liver disease worldwide. The mechanisms by which miR-34a influences NAFLD through the Sirtuin 1 (SIRT1)-related pathway were investigated herein. METHODS: Male C57BL/6 mice were injected with a miR-34a lentivirus vector inhibitor or control. HepG2 cells were transfected with a miR-34a mimic, inhibitor, SIRT1 small interfering RNA (siRNA), SIRT1 plasmid, and a negative oligonucleotide control to evaluate their role in oleic acid (OA) and excess iron-induced NAFLD. The accumulation of lipids in the mice liver and HepG2 cells was analyzed by triglyceride (TG) detection and hematoxylin and eosin (HE) staining. Additionally, the indexes of oxidative stress related to lipid metabolism were evaluated by western blotting and real-time PCR (qRT-PCR). The levels of intracellular reactive oxygen species (ROS) and mitochondrial membrane potentials were measured by flow cytometry and laser confocal microscopy, respectively. Finally, the dual luciferase reporter assay was conducted to further confirm whether SIRT1 was a direct target of miR-34a. RESULTS: Overexpression of miR-34a resulted in increased triglyceride accumulation as well as in decreased mitochondrial membrane potential and SIRT1 levels. Silencing of miR-34a increased SIRT1 expression and alleviated triglyceride accumulation in the presence of OA and iron. Additionally, miR-34a directly inhibited SIRT1 by binding to the 3'-untranslated region, as determined via the luciferase reporter assay. CONCLUSIONS: Our results support the existence of a link between the liver cell mitochondria and miR-34a/SIRT1 signaling. Potential endogenous modulators of NAFLD pathogenesis may ultimately provide new tools for therapeutic intervention.

Novel association between FOXO3 rs2232365 polymorphism and late-onset preeclampsia: a case-control candidate genetic study.

BACKGROUND: Both genetic susceptibility and dysregulated lipid metabolism are important susceptibilities to preeclampsia. In the study, we devote to investigate the associations of FOXO3 and TLR7 genetic polymorphisms with preeclampsia in a Chinese population. METHODS: This case-control study involved 335 Han Chinese pregnant women, including 177 pregnant women with preeclampsia and 158 healthy controls. The preeclampsia group was further sub-grouped into early-onset preeclampsia (EOPE, n = 70)and late-onset preeclampsia (LOPE, n = 107. Three single nucleotide polymorphisms (SNPs), including FOXO3 (rs2232365, rs3761548), and TLR7 rs3853839 were genotyped by multiplex PCR for targeted next-generation sequencing. The χ2 test and multiple interaction effect analyses were performed to determine the association of three SNPs with serum lipid levels and thyroid function in women with preeclampsia. RESULTS: The genotype (CC vs. TT + CT) distribution of rs2232365 revealed a significant association with LOPE (P = 0.004, odds ratio = 3.525 (0.95 CI: 1.498-8.164)). No significant difference was found in the genotype and allele frequencies of rs3761548 and rs3853839 between controls and cases (P > 0.05). Moreover, the genotype CT/TT of rs2232365 was significantly correlated with increased TG/HDL levels in the LOPE group (p = 0.014). CONCLUSIONS: The polymorphisms of rs2232365 are associated with the risk of LOPE and may modulate TG/HDL levels in pregnant women with LOPE.

Scattering off molecules far from equilibrium.

Pump-probe gas phase X-ray scattering experiments, enabled by the development of X-ray free electron lasers, have advanced to reveal scattering patterns of molecules far from their equilibrium geometry. While dynamic displacements reflecting the motion of wavepackets can probe deeply into the reaction dynamics, in many systems, the thermal excitation embedded in the molecules upon optical excitation and energy randomization can create systems that encompass structures far from the ground state geometry. For polyatomic molecular systems, large amplitude vibrational motions are associated with anharmonicity and shifts of interatomic distances, making analytical solutions using traditional harmonic approximations inapplicable. More generally, the interatomic distances in a polyatomic molecule are not independent and the traditional equations commonly used to interpret the data may give unphysical results. Here, we introduce a novel method based on molecular dynamic trajectories and illustrate it on two examples of hot, vibrating molecules at thermal equilibrium. When excited at 200 nm, 1,3-cyclohexadiene (CHD) relaxes on a subpicosecond time scale back to the reactant molecule, the dominant pathway, and to various forms of 1,3,5-hexatriene (HT). With internal energies of about 6 eV, the energy thermalizes quickly, leading to structure distributions that deviate significantly from their vibrationless equilibrium. The experimental and theoretical results are in excellent agreement and reveal that a significant contribution to the scattering signal arises from transition state structures near the inversion barrier of CHD. In HT, our analysis clarifies that previous inconsistent structural parameters determined by electron diffraction were artifacts that might have resulted from the use of inapplicable analytical equations.

Effects of the ionic liquid 1-hexyl-3-methylimidazolium bromide on root gravitropism in Arabidopsis seedlings.

The toxic effects of ionic liquids (ILs) have attracted increasing attention in recent years. However, the knowledge about the toxic effects of ILs on tropism in organisms remains quite limited. In this study, the effects of 1-hexyl-3-methylimidazolium bromide [C6mim]Br on root gravitropism were evaluated using Arabidopsis seedlings. Our results showed that the root growth and gravity response were significantly inhibited with increasing IL concentration. [C6mim]Br treatment affected the amount and distribution pattern of amyloplasts in root cap compared with controls. The auxin distribution marked with DR5rev::VENUS was altered in IL-treated seedlings. The signal intensity and gene expression of auxin efflux carriers PIN2 and PIN3 were obviously decreased by IL stress. Moreover, as consequences in response to gravity stimulus, the asymmetric DR5 signals in control root apex were impaired by IL treatment. The predominant PIN2 signals along the lower flank of root and PIN3 polarization in columella cells were also significantly reduced in seedlings exposed to IL. Our results suggest that the ionic liquid [C6mim]Br affects the amount and distribution of amyloplasts and disturbs the deployment of PIN2 and PIN3, thus impairing auxin flows in response to gravity stimulus and causing deficient root gravitropism in Arabidopsis seedlings.

Irradiation with low-dose gamma ray enhances tolerance to heat stress in Arabidopsis seedlings.

Gamma irradiation at low doses can stimulate the tolerance to environmental stress in plants. However, the knowledge regarding the mechanisms underlying the enhanced tolerance induced by low-dose gamma irradiation is far from fully understood. In this study, to investigate the physiological and molecular mechanisms of heat stress alleviated by low-dose gamma irradiation, the Arabidopsis seeds were exposed to a range of doses before subjected to heat treatment. Our results showed that 50-Gy gamma irradiation maximally promoted seedling growth in response to heat stress. The production rate of superoxide radical and contents of hydrogen peroxide and malondialdehyde in the seedlings irradiated with 50-Gy dose under heat stress were significantly lower than those of controls. The activities of antioxidant enzymes, glutathione (GSH) content and proline level in the gamma-irradiated seedlings were significantly increased compared with the controls. Furthermore, transcriptional expression analysis of selected genes revealed that some components related to heat tolerance were stimulated by low-dose gamma irradiation under heat shock. Our results suggest that low-dose gamma irradiation can modulate the physiological responses as well as gene expression related to heat tolerance, thus alleviating the stress damage in Arabidopsis seedlings.

Remote transfer of Gaussian quantum discord.

Quantum discord quantifies quantum correlation between quantum systems, which has potential application in quantum information processing. In this paper, we propose a scheme realizing the remote transfer of Gaussian quantum discord, in which another quantum discordant state or an Einstein-Podolsky-Rosen entangled state serves as ancillary state. The calculation shows that two independent optical modes that without direct interaction become quantum correlated after the transfer. The output Gaussian quantum discord can be higher than the initial Gaussian quantum discord when optimal gain of the classical channel and the ancillary state are chosen. The physical reason for this result comes from the fact that the quantum discord of an asymmetric Gaussian quantum discordant state can be higher than that of a symmetric one. The presented scheme has potential application in quantum information network.

EMCMDA: predicting miRNA-disease associations via efficient matrix completion.

Abundant researches have consistently illustrated the crucial role of microRNAs (miRNAs) in a wide array of essential biological processes. Furthermore, miRNAs have been validated as promising therapeutic targets for addressing complex diseases. Given the costly and time-consuming nature of traditional biological experimental validation methods, it is imperative to develop computational methods. In the work, we developed a novel approach named efficient matrix completion (EMCMDA) for predicting miRNA-disease associations. First, we calculated the similarities across multiple sources for miRNA/disease pairs and combined this information to create a holistic miRNA/disease similarity measure. Second, we utilized this biological information to create a heterogeneous network and established a target matrix derived from this network. Lastly, we framed the miRNA-disease association prediction issue as a low-rank matrix-complete issue that was addressed via minimizing matrix truncated schatten p-norm. Notably, we improved the conventional singular value contraction algorithm through using a weighted singular value contraction technique. This technique dynamically adjusts the degree of contraction based on the significance of each singular value, ensuring that the physical meaning of these singular values is fully considered. We evaluated the performance of EMCMDA by applying two distinct cross-validation experiments on two diverse databases, and the outcomes were statistically significant. In addition, we executed comprehensive case studies on two prevalent human diseases, namely lung cancer and breast cancer. Following prediction and multiple validations, it was evident that EMCMDA proficiently forecasts previously undisclosed disease-related miRNAs. These results underscore the robustness and efficacy of EMCMDA in miRNA-disease association prediction.