Effects of HCV Viral Load on Male Hormones
Sabah SaadAbdulsahib
Biomedical Engineering Department, University of Technology-Iraq, Baghdad, Iraq.
*Corresponding Author: [email protected]
ABSTRACT
The hepatitis C virus (HCV) is an infectious agent that causes human hepatitis C disease and serious life-threatening liver damage. The main objective of this research was to establish the association between the levels of plasma sex hormones and hepatitis C infection in Iraqi males. Thirty hepatitis C infected patients were included in the study sample. In addition, a control group of ten healthy peoples was created. HCV infection had been investigated and approved with TaqMan real time PCR technique. The level of hormones in the plasma of progesterone, estradiol, and testosterone had been measured via electrochemiluminescence immunoassay method usingcobas e 411 analyzers. In contrast with the controls, the findings revealed shifts in two of the three analyzed hormones within the patients. In male patients with HCV infection, the mean level of plasma testosterone and estradiol was significantly increased relative to healthy controls(P≤0.05 and P≤0.01, respectively), although there is no considerable variation between patients and controls in the mean level of plasma progesterone.The age groups have a significant effect in means of PCR viral load and progesterone hormone, while it has no significant impact in mean concentrations of hormones of testosterone and estradiol (P≤0.05). The statistical analysis for results demonstrated there is non- significant relation among HCV PCR load mean with the levels of testosterone and estradiol, though a significant correlation coefficient with progesterone level (P<0.05), the age has non-significant correlated with HCV PCR load.It was deduced that HCV disease patients have a degree of hormonal imbalance that will be explored in current study.
Keywords: Liver disease;Anabolic steroids;Estrogenic hormones; Progesterone; Sex steroids.
INTRODUCTION
The HCV infection is a major public health issue worldwide. Many cases of HCV develop into chronic hepatitis C (CHC), that may also progress to cirrhosis, hepatocellular carcinoma (HCC), and liver fibrosis.The worldwide prevalence of HCV infection is reported at over 170 million individuals, and some studies predicts that HCV infection-related mortality will continue to rise during the next two decades [1,2].The hepatitis C virus develops both chronic and acute infections. New infections with HCV are generally asymptomatic. Some peoples have acute hepatitis that does not contribute to a life-threatening illness. Approximately 30 percent of infected people spontaneously clearance the virus without treatment within six months of infection.Chronic HCV infection will evolve for the remaining 70 percent of individuals. HCV is an enveloped positive strand RNA virus that replicates readily in the liver which is included in the Flaviviridae family with in genus Hepacivirus. The genome of the HCV RNA is around 9600 long nucleotides and codes for one long polyprotein[3].TheEuropean Region and the Eastern Mediterranean Region are the most affected areas.HCV infection is also common in the general population in countries where infection management procedures are inadequate.Hepatitis C virus infection may be localized in particular populations, depending on the region. The HCV virus has several genotypes (or strains) and their spread differs by country. Nevertheless, strain distribution remains uncertain in many regions[4].Since recent HCV diseases are frequently having no symptoms of illness, less cases are detected when the infection is new. HCV infection is identified with a serological screening for anti-HCV antibodies that recognizes individuals who have been affected with the virus.A nucleic acid analysis for ribonucleic acid of HCV is required to verifyviral infection because around 30% of HCV patientsare spontaneously resolve the infection without the need for medication via a powerful immune response. They will indeed positive for immunological test even though they are no longer infected [5].Men are at dramatically greater prevalence of chronic liver disease through various etiologies of disease, particularly HCV; so, promoting the possible roles of gender-related disparitiesin risk factor exposures as well as gender-based biological differences in progression of disease [6].One of the crucial variables in the development of hepatic fibrosis due to chronic HCV infection is the male sex; therefore, woman hormones can play an important role in slowing hepatic fibrosis progression.It has also been documented that female are more likely than male to overcome
HCV in the acute phase of disease. These observations suggest the possibility that woman hormones reduce the infection with HCV, either at the stages of manufacturing of infectious virus particles,virus protein synthesis,virus RNA replication, or virus attachment/entry [7]. Antiviral drugs used to fight hepatitis C can cause sexual dysfunction (SD) and reduced libido.SD is the most common side effect of several antidepressant drugs used to curing the depression andanxietyrelated with HCV mixture treatment. SDhas been recorded to be more prevalent in patients with chronic hepatitis C than in peoples with nodefined liver disease[8].Viral etiologies of liver infections have been givenless consideration in the production of gonadal hormone malfunctions. Several investigations have reported sex hormone disorders as the cause of symptoms of liver disease.Malehypogonadism is correlated with liver disease and is characterized by low testosterone.Past research had beenconductedchanges in levels of testosterone and sex hormone-binding globulin (SHBG) in contrast with healthy controls are extrahepatic manifestations of chronic HVC[9].To date, few findings were noted regarding theimpact of HCV infection on the plasma hormone levels. Studies to investigate the impact of HCV on hormone levels are now required. Therefore, the current research was designed to establish the association between the levels of plasma sex hormones (testosterone, estradiol, and progesterone) and hepatitis C infection in some male Iraqi patients.
MATERIALS AND METHODS
A total of one hundred sixty-eight blood samples had been collected from men suspectedof hepatitis C virus infection which attending the special nursing hospital in medical city of Baghdad, for four months, starting from the first of September to the end of December 2018,ten samples were also taken from healthy individuals as control.Peoples with previously confirmed or documented infection with the HCV were the preferred chosen.Gently mix the blood after collection, by inverting the tube 8 times. By centrifugation at 1400 x g for 15 mins during four hours of blood collection, whole blood stored in EDTA tubes must be split into cellular components and plasma.The separated plasma should be transferred into a sterile polypropylene tube. Plasma had been stored at -20°C until use.
RNA extraction was carried out for all collected samples in compliance with the manufacturing instructions of the Ribo-Sorb nucleic acid extraction kit (Sacace, Italy), and stored for use at -20°C.
All RNA specimens werediagnosed byTaqMan real-time PCR (Cepheid SmartCycler, USA) assays and HCV viral load had been identified using HCV Genotype Plus Real-TM kit (Sacace, Italy) according to the protocol supplied by the kit. The findings of the PCR were analyzed with SmartCyclerDx Version 3.0b program.
Testosterone (TES), estradiol (E2), and progesterone (PROGES) plasma levels were quantitatively measured in vitro for patients and control using cobas e 411 analyzers (Roche, Germany) by electrochemiluminescence immunoassay
“ECLIA”.
To detect the effect of difference factors on study parameters, the statistical analysis system-SAS [10] program was used. In this study, the T-test was used to significantly compare means to estimate the correlation coefficient between variables.
RESULTS AND DISCUSSION
Extracted one hundred seventy-eight RNA specimens from bloodsamples of suspected cases and healthy persons were subjected to moleculardiagnosis by real time PCR to determine HCV-RNA. This assay was successful in the quantitative detection of HCV. Only 30 from 168 suspected cases were HCV-positive by real-time PCR, they were therefore regarded as presence of HCV in the specimen and male hormones investigation were performed for those 30 samples. All control samples had been given a negative result when examined by PCR.The range of ages in the studied patient range was (21-63) years with a mean age of 39.23 years, and from 20 years to 58 years as the mean age was 44.5 years for healthy control, with non-significant difference between the groups of age (Table 1).The mean results of HCV PCR load for patients were 13352976.13 ± 4950929.53 IU/ml and 0.00 ± 0.00 IU/ml for healthy control, with a highly significant difference between patient and control (P≤0.01) (Table 2).The data set in
significantly higher than healthy control (5.59 ± 0.43 ng/ml) (P≤0.05) (Figure 1). This result was in accordance with El-Serafiet al. [11] who recorded that testosterone had been significantly increased in the patients with HCV infection compared to healthy control.
Overall serum testosterone in infected males with HCV is correlated with a significantly elevated risk of both severe liver inflammatory response and chronic liver fibrosis. Testosterone could be very remarkable in the pathogenesis of chronic hepatic disease linked to HCV in men [6]. Increased concentrations of testosterone compared to individuals who achieved viral clearance of HCV were identified as active HCV infections, the association among testosterone, impaired liver function, and chronic viral infection are complicated. Effective HCV infection and associated damaged liver function are known as both risk factors for hypogonadism and changed semen development.
Hypogonadism is also a reported consequence of end phase liver illness [9]. Testosterone is mostly metabolised in the liver, and the testosterone metabolic clearance is reduced in severe liver disease [12]. An investigational research found that the HCV contributes to androgen receptor (AR) signaling rises [13]. AR is a nuclear hormone receptor expressed in the liver, the primary ligand of which is testosterone [14]. Besides controlling genes that are concerned in sexual dimorphism, AR regulates/co-regulates many other genes engaged in multiple related cellular operations for hepatic disease, which include cellular proliferation, apoptosis, differentiation, and inflammatory response [15].
Nguyen et al. [16] suggested that hepatic disease in hepatitis C infection modifies androgen status indirectly through elevated sex hormone binding globulin, checking for androgen deficiency must selectively target males with more acute liver disease or reported higher-grade fibrosis. The plasma estradiol mean level in patients with HCV (55.78 ± 3.07 Pg/ml) was significantly higher than healthy control (32.99 ± 2.05 Pg/ml) (P≤0.01) (Table 3 and Figure 2). This result was agreed with Wang et al. [17] who indicated that there were significantly higher levels of E2 in male infected with HCV than healthy males.
Farinatiet al. [18] studied type of estrogen receptor, expression of estrogen receptor, and oxidative damage to DNA in patients with chronic liver disease due to HCV, it indicated that liver positivity for an estrogen receptor variant could result in high genomic deterioration and increased rates of carcinogenesis and cytoproliferation. It was found that because of strong aromatase activity, peripheral transformation of androgens to estrogen enhanced and estrogen levels raised [19]. Conversion of adrenal androstenedione to estrone was also recorded in cirrhotic patients [20].
Iyeret al. [21] concluded sex-based variations in basal expression of estrogen receptor α (ERα) in the liver was higher in men than in women, furthermore, the increased subcellular expression of subtypes of ER present in diseased livers associates with inflammatory markers and the expression of proliferative, modulations in ER subtype expression monitored in diseased livers can affect gender-related inequality in HCV-linked pathogenesis. In the liver, estrogen and estradiol receptors defend hepatocytes from cell death, oxidative stress, and inflammatory injury, which all lead to fibrosis. Due to the overall slower development of liver disease and enhanced viral clearance in females, the risk of HCV infection is found mainly in males [22]. By restricting the aggregation of macrophages, monocytes, and suppressing the production of inflammatory cytokinesis, estradiol can serve a convenient role in chronic liver disease [23].
The mean level of plasma progesterone in male infected with HCV (0.655 ± 0.04 ng/ml) revealed non-significantly difference with healthy control (0.693 ± 0.05 ng/ml) (Table 3). This result was conflicted with the study done by Serin [24] who demonstrated that serum progesterone levels were increased in HCV liver disease patients compared to the control group. Yuan et al. [23] was observed that progesterone may reduce the E2 impacts by increasing the aggregation of inflammatory cells and the release of their cytokines by the progesterone receptor.
Table (4) showed the comparative relationship of hormonal levels and viral load in age groups. The effect of age variant was statistically non-significant in means concentrations of testosterone and estradiol, and significant effect to mean of PCR viral load and progesterone hormone (P≤0.05).The study by Okaforet al. [25] indicated that age was not correlated with the prevalence of HCV among the inspected population. Conversely, increasing age is correlated with HCV prevalence, possibly due to extended HCV infection exposure [26]. Older patients at the time of infection and with compromised immune system are at high risk of developing chronic HCV infection [27].
The statistical analysis of the findings exhibited there is non-significant negative relation among HCV PCR load mean with the levels of testosterone and estradiol, while there is a significant correlation coefficient with progesterone level (P<0.05). The age has non-significant positive correlated with HCV PCR load.
Wang et al. [17] recorded that male with increased serum testosterone levels showed inflammatory cytokines and the lowest levels of antibody after viral infection, testosterone reduces immune systems to infection with HCV. Each 1 ng/ml rise in overall serum testosterone had been linked with a 27% expanded danger of cirrhosis and a 16%
elevated risk of advanced inflammatory activity among males infected with HCV [6]. One potential reason for these 'proviral' testosterone effects is that it changes the production of lipoprotein in a way that facilitates replication of HCV [28].
Table 1.Patients and control comparison in age.
Group Mean ± SE of age (year) Patients 39.23 ±2.41
Control 40.50 ±3.98
T-test 9.658 NS
P-value 0.792
NS: Non-Significant
Table 2.Patients and control contrast in HCV-PCR.
Group No. Mean ± SE of HCV PCR Load IU/ml Patients 30 13352976.13 ±4950929.53
Control 10 0.00 ± 0.00
T-test - 327814.39 **
P-value - 0.0001
** (P≤0.01), SE: Standard Error
Figure 1. Compare between patients and control in mean of testosterone level.
Figure 2. Compare between patients and control in mean of estradiol level.
Table 3.The mean level of hormones in the plasma of the researched groups.
Group Mean ± SE
Testosterone (ng/ml) Estradiol (pg/ml) Progesterone (ng/ml) Patients 7.74 ± 0.69 55.78 ± 3.07 0.655 ± 0.04
Control 5.59 ± 0.43 32.99 ± 2.05 0.693 ± 0.05
T-test 1.419 * 11.115 ** 0.181 NS
P-value 0.0449 0.0002 0.671
* (P≤0.05), ** (P≤0.01) 7.74
5.59
0 2 4 6 8 10
Patients Control
Testosterone (ng/ml)
Groups
55.78
32.99
0 10 20 30 40 50 60
Patients Control
Estradiol (pg/ml)
Groups
Table 4. Effect of age groups in parameters study of patients.
Age group (year)
Mean ± SE HCV PCR
Load IU/ml
Testosterone (ng/ml)
Estradiol (pg/ml)
Progesterone (ng/ml) Least than 30 15871192.33
±10804297.14
7.008 ± 0.88 44.81 ± 3.64 0.531 ± 0.05
30-40 4632082.18
±4134045.95
8.23 ±1.34 51.52 ± 5.38 0.667 ± 0.09
More than 40 9363651.29
±4128580.94
6.67 ± 0.75 52.87 ± 5.09 0.756 ± 0.04
LSD value 873261.07 * 2.802 NS 14.382 NS 0.182 *
* (P≤0.05), NS: Non-Significant
CONCLUSION
The current work evaluated the potentialcorrelation between male sex hormones and HCV infected Iraqi males. It was concluded that patients with liver disease related with HCV had a degree of hormonal imbalance.The plasma testosterone andestradiol mean level was elevated significantly in meninfected by HCV in compared with healthycontrol, however, there is non-significant difference betweenpatients and control in mean level of progesterone.There is no significant correlation between mean of HCV viralload results and the means levels of testosteroneand estradiol, while there is significant correlation withprogesterone mean levels.The age has non- significant correlated with HCV PCR load.
ACKNOWLEDGEMENTS
The Department of Biomedical Engineering, University of Technology, Iraq, sponsored and supported this research.
CONFLICT OF INTEREST
The author declare that there is no conflict of interests regarding the publication of this manuscript.
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