Nasopharyngeal　carcinoma　(NPC)　is　an　aggressive　head　and　neck　malignancy　originating　from　the　epithelial　lining　of　the　nasopharyngeal　mucosa,　which　is　closely　associated　with　the　squamous　metaplasia　of　pseudostratified　ciliated　columnar　epithelium　(PCCE).　Multiphoton　microscopy　(MPM)　is　an　advanced　biomedical　imaging　technique　based　on　nonlinear　optical　effects,　such　as　two-photon　excited　fluorescence　(TPEF)　and　second　harmonic　generation　(SHG).　It　offers　high-resolution,　deep-tissue　imaging　capabilities　without　the　need　for　exogenous　contrast　agents.　In　this　study,　MPM　in　combination　with　spectroscopic　imaging,　was　employed　to　image　nasopharyngeal　tissue　samples,　to　investigate　the　structural　changes　from　normal　pseudostratified　ciliated　columnar　epithelium　to　squamous　metaplasia.　The　results　demonstrated　that　MPM　enabled　high-resolution　visualization　of　nasopharyngeal　microstructures　without　external　contrast　agents,　and　spectroscopic　analysis　effectively　monitored　the　dynamic　structural　changes　associated　with　epithelial　squamous　metaplasia.　This　study　provided　novel　insights　into　the　microscopic　structural　alterations　during　squamous　metaplasia　in　NPC,　offering　potential　applications　in　early　diagnosis　and　pathological　research　of　NPC.　©　2025　SPIE.