The aggregation of Medin is closely related to the arterial wall degeneration and cerebrovascular dysfunction. In patients with vascular dementia or Alzheimer’s disease, the concentration of medin in cerebral arterioles increases, and Medin is co-localized with vascular amyloid-β (Aβ) deposits. Previous study demonstrates that Medin interacts directly with Aβ, forming heterologous fibrils with Aβ and promoting Aβ aggregation. However, the basic mechanism of the co-aggregation between Medin and Aβ remains largely elusive. Here, we explore the interactions and conformational ensembles of Aβ₄₂/Medin trimers in different peptide environments (self-aggregation vs. co-aggregation) by performing all-atom replica exchange molecular dynamic simulation on Aβ₄₂/Medin homotrimers and Aβ₄₂-Medin heterotrimer with an accumulated simulation time of 72 μs. Our results reveal that Aβ₄₂ exhibits higher affinity with Medin, and Aβ₄₂ and Medin have similar molecular recognition sites in self-aggregation and co-aggregation. The N-terminus of Aβ₄₂ and the C-terminus of Medin play critical roles in Aβ₄₂-Medin cross-talk. More importantly, co-aggregation significantly changes the interaction strength, binding patterns and structural characteristics of Aβ₄₂ and Medin. Intermolecular interactions of Aβ₄₂ trimers are relatively weak among three trimers, and the binding sites are concentrated between N- and N-termini, between N- and C-termini, and between C- and C-termini of Aβ₄₂. In contrast, intermolecular interactions of Medin trimers are the strongest, and the binding sites are widely and uniformly distributed in Medin peptides. Intermolecular interactions of Aβ₄₂ in Aβ₄₂-Medin heterotrimer decrease compared with those of Aβ₄₂ trimers, only the binding of the hydrophobic core regions (¹⁶KLVFFA²¹) is retained and other regions of Aβ₄₂ gain increase flexibility. Two-dimensional free energy landscapes reveal distinct conformational diversities between the homo- and heterotrimers, with the order of diversity being Medin/Aβ42-Medin trimers > Aβ₄₂ trimers. The R_g of Aβ₄₂ trimers is smaller than those of the other two trimers, implying that Aβ₄₂ trimers possess a more compact structure, whereas Medin/Aβ₄₂-Medin trimers exhibit a relatively loose conformation. The Aβ₄₂ trimers possess the highest β content whereas Medin trimers exhibit the lowest β probability. It is found that Aβ₄₂-Medin co-aggregation induces Medin to form more β-structures with longer lengths and fewer helices, while promoting Aβ₄₂ to form more helices and fewer β-structures. High β-propensity regions of Medin in heterotrimers shift towards the C-terminus of Medin, suggesting that Medin utilizes its C-terminal β region as a core motif to drive its co-aggregation with Aβ₄₂. These results elucidate the detailed influences of co-aggregation on the interactions and conformations of Aβ₄₂ and Medin. This work provides key insights into the molecular mechanism of Aβ₄₂-Medin co-aggregation and the pathological mechanisms of cross-linking between related diseases.