The attack on the militia positions were described as a "defensive" measure by Washington, after fighters - believed to be linked to Iran - attacked a US-backed anti-Islamic State group coalition in eastern Deir az-Zour province.
Scores were killed in the US airstrikes, after the pro-regime militias opened fire on an oil installation manned by the pro-Washington Syrian Democratic Forces (SDF).
"We estimate more than 100 Syrian pro-regime forces were killed while engaging SDF and coalition forces," a US military official told AFP.
Syria's regime and pro-Assad Russia were furious with the strikes.
"[The attack] represents a war crime and a crime against humanity," the Syrian regime said in a letter to United Nations Security Council and the body's secretary general.
"[It represents] dirty US intentions against Syria's sovereignty and territorial unity," it added.
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Russia said the air strikes only wounded 25 fighters, but claimed the measure points to US designs for the resource rich area.
"[It] again showed that the US is maintaining its illegal presence in Syria not to fight the Islamic State group, but to seize and hold Syrian economic assets," Moscow claimed.
The US has backed the Syrian Democratic Forces in their fight against the Islamic State group in northern and eastern Syria.
Kurdish and Arab fighters from the force have led the campaign to defeat IS in Syria, capturing the group's stronghold of Raqqa in October.
Pro-regime and Russian forces have concentrated on defeating Syrian rebels over the past three years.
Washington and the Syrian opposition said they have only put minimum effort into fighting IS and waited until the group were severely weakened by the Kurdish-Arab force before launching an offensive.
Since then, Syrian regime forces - backed by Russia and Iran - have tried to carve out as much of oil and gas-rich Deir az-Zour province as possible.
The US has concentrated its attacks on IS and al-Qaeda positions, but has also carried out periodic strikes on Iranian-sponsored militias in southern Syria.