In 2002, the World Health Organization declared that zinc deficiency stands as one of the foremost risk factors contributing to morbidity and mortality, particularly in developing countries (Ozeki et al. 2020). Poor survival in chronic viral patients with early HCC was significantly associated with Zn deficiency (Kim et al. 2020).

Chronic liver disease (CLD) is among the conditions associated with diseases that can lead to zinc deficiency in adults (Ozeki et al. 2020).

Zinc deficiency in chronic liver diseases arises from a combination of factors, including a reduced capacity to synthesize albumin, malabsorption of zinc from the intestine, and an elevated excretion of zinc in the urine (Himoto et al. 2020; Ozeki et al. 2020; Grüngreiff et al. 2016; Fathi et al. 2020).

Previously reported RCTs demonstrated the potential of zinc supplementation to ameliorate hepatic encephalopathy-related clinical symptoms, as well as to enhance clinical parameters (such as blood albumin, ammonia levels, and Child–Pugh score) or to stop the evolution of the disease (e.g., progression to HCC) (Diglio et al. 2020; Shipley et al. 2019).

Previous clinical studies have shown that polaprezinc (a protector of the gastric mucosa) has been recently demonstrated to be an inhibitor of liver fibrosis in a mouse model suggesting that it may have antifibrotic effect in people suffering from autoimmune hepatitis and liver cirrhosis as it may affect collagen degradation through deactivation of hepatic stellate cells (HSCs) by inhibiting both cell behavior transitions and essential gene production and also modulation of the activity of matrix metalloproteinases (MMPs) and their tissue inhibitors of metalloproteinase (TIMs) (Ye et al. 2017; Moriya et al. 2021; Takahashi et al. 2007).

We aimed in the present study to focus on the potential antifibrotic impact of zinc sulfate in chronic HCV patients with mild and moderate fibrosis receiving direct-acting anti-viral therapy.

Zinc supplements influence hepatic fibrosis by decreasing the elevated fibrogenic activity in late stages of liver injury and increasing hepatic collagenolysis which is suppressed in early stages of liver fibrosis (Giménez et al. 1994).

Zinc supplementation results in a significant reduction in collagen deposition in the liver. Furthermore, zinc suppresses gene expression of α-SMA and collagen I and enhances the capacity of collagen degradation as determined by the increased activity of total collagenases and elevated mRNA and protein levels of MMP13, so zinc supplementation suppresses liver fibrosis through both inhibiting collagen production and enhancing collagen degradation (Shi et al. 2015).

Zinc supplementation attenuated the increase in factors known to be associated with hepatic apoptosis (Zhou et al. 2008; Kang et al. 2008).

Zinc deficiency causes a decline in the activity of collagenase that leads to liver fibrosis (Stamoulis et al. 2007; Seltzer et al. 1977). Moreover, zinc exerts membrane-stabilizing activity on liver lysosomes and has cytoprotective activities that protect hepatocytes from oxidative stress (Zhou et al. 2005; Stamoulis et al. 2007; Kono et al. 2012).

In the present study we used the standard dose of zinc sulfate that provides the patients with 50 mg of elemental zinc, which represent the daily requirement of the human body for 3 months which matches some previous clinical studies (Somi et al. 2012; Mohammad et al. 2012; Shirahashi et al. 2021; Hosui et al. 2021).

Despite being one of the most accurate ways to evaluate the condition of the liver, liver biopsies are an intrusive procedure and may lead to severe complications, bleeding and hemodynamic instability (Chowdhury et al. 2023; Boyd et al. 2020).

Consequently, we employed safe and noninvasive techniques to measure liver fibrosis in this study, such as measuring serum levels of liver fibrosis biomarkers (i.e., HA, FN and TGF-β1) and calculating FIB-4 and APRI scores. It has been shown that HA, FN and TGF-β1 are the most useful diagnostic and prognostic noninvasive biomarkers for hepatic fibrosis (Ghafar et al. 2019; Boyd et al. 2020; Ehsan et al. 2021; Rewisha et al. 2021; Dewidar et al. 2019). Additionally, it was observed that the FIB-4 and APRI scores had a diagnostic precision for determining the liver fibrosis stage (Itakura et al. 2021).

In the current study, the administration of zinc sulfate showed a significant improvement in serum zinc level and a remarkable decline in the liver fibrosis index (APRI score) and biomarkers (HA and FN) when compared to the control group. Bilirubin level significantly decreased within zinc group and between the two study groups after treatment as revealed in previous studies which indicate the positive effect of zinc on the liver (Hiraoka et al. 2020).

The phases of liver fibrosis range from non-cirrhotic stages (F0–F3) to cirrhotic stage (F4). Fibrosis, characterized by the replacement of parenchyma with a collagenous scar due to repetitive liver hits, is influenced by different etiologic agents including diabetes, alcohol intake, metabolic syndrome, viral infections like HCV and many other diseases (Shirahashi et al. 2021; Masuzaki et al. 2020; Grüngreiff et al. 2016).

Opposing the traditional view that it is a unidirectional process, several research studies have demonstrated liver fibrosis's dynamic nature and potential for reversibility. Preventing, decreasing and reversing the progression of fibrosis to cirrhosis and its related problems is the main goal of current and future therapy for chronic liver disease, this will lessen the need for liver transplantation (Himoto et al. 2020; Iritani et al. 2022).

The liver plays a crucial role in maintaining the body's zinc homeostasis. Numerous chronic liver diseases, such as nonalcoholic fatty liver disease, chronic viral hepatitis, and liver cirrhosis, have been linked to zinc deficiency. Such deficiency can lead to the impairment of multiple hepatic functions. Furthermore, studies have indicated a correlation between zinc deficiency and liver fibrosis in chronic HCV patients (Omran et al. 2017).

Liver fibrosis occurs due overproduction of extracellular matrix (ECM) which consists of substances including glycoproteins, collagen, proteoglycans, and glycosaminoglycans.

Increased production and accumulation of extracellular matrix in liver tissues affect the normal liver cell functions and increases the progression of fibrosis. The concentrations of ECM in blood serve as biomarkers of fibrotic diseases (Matsuda et al. 2020).

HCV replication and proliferation activate immune cells in the liver which are for example, Kupffer cells, macrophages and natural killer cells that leads to activation of hepatic stellate cells (HSC) that secrete fibrillar collagens (Shipley et al. 2019). In hepatic fibrosis, liver sinusoidal endothelial cells are also beneficial and essential cells. These cells have the capacity to activate HSC and produce fibronectin in the very early stage of liver damage.

Fibronectin is non-collagenous glycoprotein fundamental constituent of ECM and is considered a reliable marker of liver fibrosis (Acharya et al. 2021).

In our study, fibronectin level was significantly higher in the control group; however, there was no significant difference in fibronectin level in the zinc group. In addition, administration of zinc sulfate caused a significant decrease in serum fibronectin levels in zinc group compared to the control which supports a prior study that found zinc reduces the gene expression of liver fibrotic markers, such as fibronectin in vitro. (Ye et al. 2017; Shan et al. 2023).

The most important and fundamental glycosaminoglycan of extracellular matrix (ECM) is hyaluronic acid (HA), which is primarily produced by hepatic stellate cells and broken down by sinusoidal endothelial cells. According to previous studies, it may correlate with the histological phases of liver fibrosis in chronic liver disorders (Takahashi et al. 2007).

In our clinical research, zinc sulfate caused a significant decrease in serum hyaluronic acid levels in zinc group compared to the control and compared to their baseline value, which agrees with a previous study that zinc level is negatively correlated with hyaluronic acid level in patients with autoimmune hepatitis (Shan et al. 2023).

This may be due to the ability of zinc supplementation to inhibit of hepatic stellate cells activation, as reported in prior animal and in vitro studies (Ye et al. 2017; Xie et al. 2021).

TGF-β1 is a profibrotic cytokine known for stimulating the cellular synthesis and deposition of molecules within the extracellular matrix, including collagen types I, II, IV, and elastin. HSCs are the primary source of TGF-β1 although other cell types, such as macrophages, liver cells, and thrombocytes, also have the capacity to secrete this cytokine. The multifaceted involvement of various cell types underscores the intricate role of TGF-β1 in the fibrotic processes within the liver (Dewidar et al. 2019).

Additionally, zinc sulfate caused a non-significant decrease in serum TGF-β1 levels in the zinc group compared to the control which may be due to our patients being in early stages of liver fibrosis (F0, F1, F2) and the relatively small sample size.

The non-significant results of FIB-4 between the two groups indicate that the degree of liver fibrosis was minimal and varied between F0, F1, and F2, or may be caused by the limited number of patients.

Nevertheless, the notable decline in fibronectin and hyaluronic acid serum levels and significant negative correlations between zinc level and each of hyaluronic acid and TGF-B levels reflect the role of zinc supplementation to act against the mechanism of liver fibrosis and may represent what happened in the early stages of liver fibrosis in HCV.

Hence, our conclusions appear to be consistent with a few other reports which concluded that zinc concentrations reduced significantly with the progression of liver fibrosis and considered oral zinc supplementation to be beneficial in treating chronic hepatitis C (Hosui et al. 2018, 2021; Nishikawa et al. 2020; Kiouri et al. 2023).

One of the study's highlights is that it is the first clinical research focused on comparing the effectiveness of oral zinc sulfate tablets for liver fibrosis in Egyptian patients with hepatitis C virus who are receiving the DAAs combination of sofosbuvir 400 mg and daclatasvir 60 mg.

While our clinical research yielded beneficial results, it is important to acknowledge certain limitations. These include the study's open-label design, a single dose of 50 mg elemental zinc and a somewhat small sample size. Therefore, further larger-scale research with other doses of zinc and longer duration is necessary to confirm and expand on our findings.