Heat-induced decreases in bovine blastocyst development have been related to reductions in latent matrix metallopeptidase-9 (proMMP9) production from maturing cumulus-oocyte complexes. Elevated intrafollicular proMMP9 levels at the time of oocyte retrieval have been positively related to pregnancy following human IVF. Thus, we hypothesized that heat-induced reductions in proMMP9 levels during oocyte maturation may be responsible for decreased blastocyst development. As a first step towards testing this hypothesis, bovine cumulus-oocyte complexes were matured at 38.5°C for 24 h with 0 or 300 ng/mL recombinant human proMMP9 (rhMMP9) added at 0 h of in vitro maturation (hIVM). No differences were found in ability of oocytes to cleave or form blastocyst-stage embryos after IVF. In a second study, cumulus-oocyte complexes were matured at 38.5 or 41.0°C (first 12 h only, then transferred to 38.5°C). At 12 hIVM, 0 or 300 ng/mL of rhMMP9 was added. Heat stress exposure decreased 24 hIVM proMMP9 levels in 0 (P = 0.006) but not 300 ng/mL groups and elevated progesterone levels most when 300 ng/mL rhMMP9 was added (P = 0.0002). Heat stress exposure did not affect ability of oocytes to cleave but reduced blastocyst development (P = 0.006). Independent of maturation temperature, addition of rhMMP9 decreased cleavage (P = 0.02) and blastocyst development (P = 0.08). In a third study, 0, 30 or 300 ng/mL rhMMP9 was added at 18 hIVM to cumulus-oocyte complexes matured at 38.5 or 41.0°C (first 12 h only, then transferred to 38.5°C). Heat stress exposure decreased 24 hIVM proMMP9 levels in 0 (P = 0.007) and 30 (P = 0.04) but not 300 ng/mL groups and increased progesterone levels in 0 and 300 but not 30 ng/mL rhMMP9 groups (P = 0.039). Heat stress exposure decreased cleavage (P \u3c 0.0001) and blastocyst development (P \u3c 0.0001). Independent of maturation temperature, addition of rhMMP9 did not alter cleavage but decreased blastocyst development (P = 0.02). In summary, addition of rhMMP9 at evaluated doses and times during IVM did not restore development of heat-stressed oocytes. Addition of 30 or 300 ng/mL rhMMP9 after 12 hIVM, regardless of maturation temperature, was detrimental to development
