Abstract: In order to fulfill the technical requirements of a high-efficiency low-emissions off-road horizontal diesel engine and design a rational cooling system, combined with the cooling water flow test, a numerical simulation model of the coolant flow was built by using three-dimensional Computational Fluid Dynamics (CFD) simulation method and was verified by the comparison between the measurement data and the CFD simulation data. The impact of cooling water jacket structure on coolant flow field in 2D25 horizontal diesel engine with a forced-cooling closed-loop cooling system, and the cooling water jacket structure was finally optimized. The results indicated that the rational design of the water inlet holes in the cylinder block can improve the coolant flow characteristics and cooling effect in the water jacket, reduce the flow losses and the water pump power loss. Rational design of the water inlet holes in the cylinder head can greatly increase the overall coolant flow velocity in the cylinder head water jacket, reduce cylinder-to-cylinder variation in cooling, improve the cooling in the critical valve bridge area in the cylinder head to sustain high thermal loading, and enhance the effectiveness of cooling at the exhaust port side. After optimization, the overall spatial average velocity and the overall spatial average heat transfer coefficient of the coolant in the water jacket increased by 40% and 41.7% respectively compared to the original design. There are no large vortices any more in the shared water chamber and the upper region of each cylinder water jacket. The flow velocity and heat transfer in the critical valve bridge area in the cylinder head were significantly enhanced, and the uniformity of cylinder-to-cylinder was improved.