Nene played only 1 minute, 43 seconds before suffering the injury. Houston coach Mike D'Antoni opted to play power forward Ryan Anderson as the backup center after Nene's departure.
"He pulled his groin pretty good," D'Antoni said of Nene, who was taken to an area hospital for an MRI. "We'll see. I don't have more than that."
Nene entered Sunday's game against the San Antonio Spurs averaging 10.8 points and 5.3 rebounds in 19.9 minutes per game during the playoffs.
With Houston's 125-104 win, the series returns to San Antonio tied 2-2.
ESPN's Michael C. Wright contributed to this report.