A hydroxyproline assay was performed to quantify collagen content in both the PSF and PIF, as described in previous studies.12, 24 The samples were hydrolyzed overnight in 0.5 ml of 6 M hydrochloric acid at 105 °C. 10 μl of hydrolysate were mixed with 150 μl isopropanol followed by 75 μl of 1.4% chloramine‐T (Sigma, St. Louis, Missouri) in citrate buffer and oxidized at room temperature for 10 minutes. The samples were then mixed with 1 ml of a 3:13 solution of Ehrlich reagent [1.5 g of 4‐(dimethylamino)benzaldehyde (Sigma), 5 ml ethanol, and 337 μl sulfuric acid] to isopropanol and incubated for 45 minutes at 55 °C. Quantification was determined by extinction measurement of the resulting solution at 558 nm. A standard curve (0–1,000 μM, trans‐4‐hydroxy‐l‐proline; Sigma) was included in each assay. Results are reported as micrograms of hydroxyproline per milligram of tissue wet weight.