Gallbladder disease, presenting commonly as gallstones and inflammation (cholelithiasis and cholecystitis), represents a marked global health burden and is a leading cause of hospital admissions related to gastrointestinal diseases [23]. Laparoscopic cholecystectomy (LC) is known to be the gold standard and one of the most frequently performed abdominal surgeries for symptomatic gallbladder disease worldwide [24]. Although laparoscopic cholecystectomy (LC) has several benefits, it is associated with a considerable risk of iatrogenic bile duct injuries (BDI), with reported incidences ranging from 0.08% to 1.5% [25]. These injuries are a major issue due to their association with notable morbidity and mortality, decreased quality of life, and substantial healthcare costs, including complications such as biliary stricture, sepsis, and recurrent cholangitis [25]. In response to these challenges, near-infrared cholangiography (NIRC) with indocyanine green (ICG) has emerged as a promising solution to enhance intraoperative visualization of the biliary tree, achieve a critical view of safety, and reduce complications like biliary injuries [26].

Liu et al. [27] is an animal study that was implemented on seven swine. This study found that near-infrared fluorescence cholecysto-cholangiography using a direct intragallbladder injection of ICG dye provides more real-time guidance to identify biliary structures in comparison with intravenous injection of the dye. Compared to systemic injection, this method offers a faster and better contrasted enhancement of Calot’s triangle and the gallbladder itself.

A limited number of human studies have compared the two injection techniques, and their results have also been conflicting. Therefore, this meta-analysis aims to compare the two injection techniques in terms of different biliary tree structure visualization, operative time, and postoperative complications.

This meta-analysis demonstrated no statistically significant differences between the two injection methods for cystic duct visualization, common bile duct visualization, and common hepatic duct visualization. However, statistically significant differences were observed for liver visualization, favoring the intravenous group compared with the intrabiliary group (RR = 0.08, 95% CI 0.03–0.21, P < 0.00001), and for operative time, which was longer in the intrabiliary injection group compared with the intravenous group (MD = 8.80, 95% CI 1.05–16.56, P = 0.03). Bile duct injury was reported in three studies, with zero events in both groups, precluding calculation of a comparative effect.Given the small sample size, few outcome events, and substantial clinical and methodological heterogeneity across the included studies, these findings should be interpreted with caution and considered hypothesis-generating only. Furthermore, the GRADE assessment rated the certainty of evidence for all primary outcomes as very low due to study limitations, inconsistency, and imprecision; therefore, no conclusions regarding clinical superiority or practice-changing implications can be drawn.The reason for the increased liver fluorescence in the intravenous injection group is that intravenous ICG injection is delivered systemically, allowing hepatic uptake and parenchymal accumulation of the dye before biliary excretion. In contrast, intrabiliary injection introduces the dye directly into the biliary system, bypassing the hepatic parenchyma, and therefore yields minimal liver fluorescence. This increase in liver fluorescence has both advantages and disadvantages. One of the advantages is that it indicates the dye has reached the normal biliary ducts, enhancing the visualization of the biliary anatomy and helping to reduce the risk of biliary tree injury. On the other hand, one of the disadvantages is that excessive liver fluorescence may create background noise, making it more difficult to identify smaller ducts, particularly in cases with inflammation or adhesions [14, 20].

The increase in operative time with intrabiliary injection is mainly due to the additional technical steps required during surgery. These include accessing and puncturing the gallbladder or cystic duct to inject the dye, confirming correct catheter or needle placement, and managing potential dye leakage, which may require extra suction or irrigation. Intrabiliary injection also demands more preparation and carries a higher risk of technical difficulties compared to intravenous injection [14].

In the subgroup analyses, neither study design group showed a meaningful difference in CD visualization before or after dissection between the two injection methods, and findings were consistent with the overall effect. For operative time, although each subgroup alone showed no significant difference, the pooled analysis indicated a slight increase in operative time with intrabiliary injection.

CBD visualization before dissection of Calot’s triangle, CBD visualization not determined before or after dissection, and CHD visualization before dissection of Calot’s triangle demonstrated risk ratios below 1, favoring intravenous injection; although the confidence intervals crossed the line of no effect, resulting in no significant pooled difference.

For CBD and CHD visualization after dissection, the two studies showed opposite directions of effect; however, both were non-significant. In contrast, liver fluorescence consistently favored intravenous injection, with both studies showing significant risk ratios below 1 and confidence intervals that did not cross the line of no effect. Nevertheless, the robustness of these subgroup findings is limited by the very low certainty of evidence as per GRADE, restricting the confidence with which these differences can be generalized to broader clinical practice.

One of the strengths of this meta-analysis is that it includes all types of study designs—covering randomized controlled trials, retrospective studies, and case-control studies—with a total of four studies involving a sample size of 178 patients, aiming to minimize bias through comprehensive study inclusion.

However, the limitations of this meta-analysis must be acknowledged, including the inherent biases associated with the inclusion of retrospective studies, potential heterogeneity among the included studies. Second, the limited number of included studies with small sample sizes makes the heterogeneity statistics and subgroup effects unstable when the number of studies is small [1, 2]. Additionally, we did not include grey literature such as protocols from clinical trial registries or sources indexed through Google Scholar. Furthermore, although our protocol restricted inclusion to English-language studies, this criterion did not ultimately impact study selection, as no eligible non-English articles were identified during screening. Collectively, these limitations contributed to the very low GRADE certainty ratings, emphasizing the need for larger, high-quality trials to increase confidence in the observed effects.

In conclusion, this systematic review and meta-analysis was conducted to explore the hypothesis that the route of indocyanine green (ICG) injection may influence biliary anatomy visualization during laparoscopic cholecystectomy. The findings indicate that intrabiliary and intravenous ICG injection techniques provide comparable visualization of the biliary anatomy. Differences were observed in liver fluorescence characteristics and operative time between the two injection routes. Notably, lower liver fluorescence associated with intrabiliary injection may reduce background hepatic signal, potentially improving visual clarity of the biliary structures and facilitating more precise anatomical identification during surgery. Additionally, operative time was observed to be slightly longer with intrabiliary injection. The outcome “bile duct injury” remains non-informative, as no events occurred in either group, precluding any comparative inference. These observations should be interpreted with caution and do not imply clinical superiority.

According to the GRADE assessment, the certainty of evidence for all primary outcomes was rated as very low due to study limitations, inconsistency, imprecision, small sample sizes, few outcome events, and heterogeneity across study designs. Therefore, the present findings should be considered hypothesis-generating only. Larger, well-designed, adequately powered randomized controlled trials are required to determine whether these observed differences translate into clinically meaningful benefits.