With continuous urbanization, human activities have left considerable impacts on the ecology. Therefore, it is necessary to perform timely and objective monitoring and evaluation of the ecology. With the basin of three highland lakes (Fuxian Lake, Xinyun Lake, and Qilu Lake) in Yunnan Province as the study case, four indices, i.e., the Normalized Difference Vegetation Index (NDVI), the Wet Index (WET), the Normalized Differential Build-Up And Bare Soil Index (NDBSI), and the Land Surface Temperature (LST), which indicate, respectively, greenness, humidity, dryness, and heat of the study area, were extracted. On the basis of five sets of terrestrial images of the areas around the three lakes from 2001 to 2021, principal component analysis (PCA) was performed on these four indices; the more informative principal component contribution was selected as the weight to establish a remote sensing ecological index (RSEI) evaluation model to evaluate the ecological environment quality of the study area; the Mann–Kendall test combined with Sen’s slope (Sen + MK) and the Hurst exponent were employed to explore the ecological conditions and development trends of the “three-lake” basin. The results showed that the ecological quality of the study area improved and then deteriorated from 2001 to 2021. The ecological quality classes in the study area were fair, medium, and good. The ecological quality has been greatly improved, but poor ecological quality was still observed in some regions such as Chengjiang. Eighty-eight percent of the study area witnessed a stable trend in the ecological quality over the 20 years; in 2021, the area of built-up land with fair and poor ecological quality reached 140.97 km2, which occupies 68.1% of the total area under the same land use type. Analysis shows that urban area expansion and human activities have exacerbated ecological problems of towns and built-up land in the study area. In the selected indicators, both greenness and humidity are positive indicators to ecological quality, and the R2 value of the two in 5-year regression was both greater than 0.99, which validated the reliability of the selected model indicators. The research findings are expected to provide a basis for scientific ecological planning and restoration of lake basins.