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Journal of Clinical and Experimental Hepatology logoLink to Journal of Clinical and Experimental Hepatology
. 2021 Feb 4;11(6):691–699. doi: 10.1016/j.jceh.2021.01.006

Gilbert's Syndrome, Bilirubin Level and UGT1A1∗28 Genotype in Men of North-West Region of Russia

Andrei Ivanov ∗,, Elena Semenova
PMCID: PMC8617539  PMID: 34866848

Abstract

Background/Objectives

Gilbert's syndrome (GS) is a hereditary pathology that affects approximately 10% of the world's population. In most cases, GS is associated with the UGT1A1∗28 polymorphism of UGT1A1 gene coding the enzyme bilirubin uridine diphosphate glucuronosyltransferase (UGT-1A) which plays a key role in the bilirubin metabolism. The presence of an additional TA repeat in the TATA box of the UGT1A1 gene promoter (the allelic variant of 7TA, abbreviated as UGT1A1∗28) leads to a significant decrease in the enzymatic activity of UGT-1A in the liver and to decrease in glucuronidation process as a consequence. The aim of the study is to estimate the prevalence of the 6TA/6TA, 6TA/7TA, and 7TA/7TA genotypes of UGT1A1 promoter and to analyze the effect of these variants on bilirubin levels in healthy men in North-West Russia and patients with a clinical diagnosis of GS.

Methods

Genotyping of the UGT1A1 ∗28 (rs8175347) polymorphism was carried out by real-time PCR.

Results

The results obtained indicate an increased probability of GS developing in residents of the North-West region of Russia compared with other representatives of the Caucasians.

Conclusions

Despite the fact that the level of serum bilirubin increases with the rise in the number of additional TA dinucleotides in the UGT1A1 gene promoter tests of clinical manifestations only (jaundice, fatigue, sleep disturbances, nausea, belching, and so on) and increased bilirubin levels in patients with normal liver function do not allow unequivocally diagnose GS. UGT1A1∗28 genotyping should be used as a prognostic risk factor for such pathology development.

Keywords: Gilbert's syndrome, hyperbilirubinemia, UGT1A1∗28 polymorphism

Аbbreviations: ATV, atazanavir (protease inhibitors); BaP, benz(a)pyrene; CHD, coronary heart disease; CVD, cardiovascular disease; GS, Gilbert's syndrome; UDP, uridine diphosphate; UGT, enzyme bilirubin uridine diphosphate glucuronosyltransferase; UGT-1A, uridine diphosphate glucuronosyltransferase isoform 1А


Gilbert's syndrome (GS) is a common hereditary pathology characterized by unconjugated hyperbilirubinemia in the absence of bilirubinuria and obvious hemolysis.1 From the hepatology point of view GS is considered benign since it does not lead to liver dysfunction or fibrosis.2 With GS, the processes of bilirubin conjugation in hepatocytes are disrupted due to the partial or complete absence of the bilirubin uridine diphosphate glucuronosyltransferase (UGT-1A) enzyme activity.3,4

The prevalence of hereditary unconjugated hyperbilirubinemia due to a decrease in the enzymatic activity of UGT-1A in hepatocytes varies widely and depends on ethnicity amounting up to 15–25% for Africans and demonstrating lower rates (from 0-5% to 5–15%) for Asian and European populations, respectively.4,5 The diagnosis of GS is made by detecting an elevated level of bilirubin in the blood (more than 21 мmol/L) with normal liver function. GS is considered as a multifactorial disorder.6 Despite the fact that it is a congenital disorder GS may remain undiagnosed until the end of adolescence.7 Another characteristic feature of this pathology is the connection with sex. Despite the significantly stronger clinical manifestations in men, women also suffer from GS especially in the presence of provoking factors (ratio 2–7 to 1). Specifically the prevalence of GS is 12.4% among Caucasian men and 4.8% among Caucasian women.8 Apparently the higher GS prevalence in men can be explained by two reasons: increased compared with women production of bilirubin and inhibition of bilirubin glucuronidation by androgens.7

Today more than 110 UGT1A1 gene variants have been described differing both in the coding sequence and in the promoter region. They occur with different frequencies in different ethnic groups.7 GS is often associated with a functional nucleotide polymorphism of the UGT1A1 gene promoter which provides a reduced enzymatic activity of UGT-1A and as consequence unconjugated hyperbilirubinemia.9 Polymorphism is come out in TA repeats number difference in the UGT1A1 promoter. A feature of this genetic model is the UGT1A1 gene expression regulation using the TATA box. Several TA dinucleotide repeats are added in the UGT1A1 gene promoter to the beginning of the TATA box: А(ТА)nТАА.9 It is considered that the normal (wild type) genotype is six repeats presence (n = 6; A(TA)6TAA) which ensures the optimal level of gene expression. Most cases of GS are associated with one extra TA insert in the TATA box.10 Such genotype (n = 7; A(TA)7TAA) is known as genetic cause of altered plasma bilirubin levels in the adult population accounting for 18% of all the UGT1A1 gene major variations in Europeans.11 The allelic variant with seven TA repeats received the abbreviation UGT1A1∗28.12 This polymorphism significantly reduces UGT1A1 expression7,13,14,15 and leads to increase of the unconjugated bilirubin level.7,12 Polymorphism ∗28 reported in 26–31% of Europeans and 42–56% of African Americans.7,16 In Asian populations and among residents of the Pacific Islands, the allelic variant UGT1A1∗28 occurs with much less frequently: 9–16% and 4%, respectively.13,16 About 20% of the white population of the Earth is homozygous carriers of the UGT1A1∗28 allele.2

Unfortunately, despite a significant number of publications devoted to both the clinical manifestations of GS and the genetics of UGT1A1, many biological features of this pathology remain unclear.17

Considering the higher prevalence of GS in men as well as the fact that in the vast majority cases described in the scientific literature clinical hyperbilirubinemia in patients with GS is associated with the mutant allele UGT1A1∗28 we focused in presented study on the UGT1A1∗28 polymorphism (rs8175347) genotyping in men of the North-West of Russia. The aim of the study was to assess the frequency of the allelic variant UGT1A1∗28 in men with a clinical diagnosis of GS and in the control group of healthy blood donors and to compare the specific genotype with the plasma bilirubin concentration in both groups. This case-control study was carried out in a large medical center – Saint-Petersburg State University Hospital (Russia). The results obtained indicate an extremely high prevalence of the pathological allele UGT1A1∗28 in male population of North-West Russia and confirm the relationship between UGT1A1∗28 polymorphism and hyperbilirubinemia in patients with GS.

Methods

Patients

The project involved 198 Caucasian men permanently residing in the North-West region of the Russian Federation. The patients group consisted of 124 men of different age 26–56 years who for a long time demonstrated obvious clinical manifestations of hereditary hyperbilirubinemia aimed at genetic confirmation of the Gilbert's syndrome diagnosis in the Saint-Petersburg State University Hospital. The average age of patients with GS was 31.1 ± 19.15 years.

The control group included 74 healthy men with no history of diseases associated with elevated bilirubin levels complaints. The age of the study participants in this group was 30–39 years; the average age was 33.92 ± 2.87 years.

The study was conducted in accordance with the Declaration of Helsinki by the World Medical Association. All study participants completed and signed an informed consent to participate in this research project and to publish the results. All clinical data were depersonalized. The study protocol was approved by the Ethics Board of Saint-Petersburg State University Hospital.

Biochemical testing

Blood sampling was performed on an empty stomach by venipuncture using vacuum tubes Lind-Vac, Estonia.

Bilirubin level definition in the blood was carried out on an automatic biochemical analyzer OlympusAU-680 (Beckman Coulter, USA) according to the instruction.

Genetic testing

DNA isolation from peripheral blood leukocytes was carried out by the method of binding on magnetized silica at a robotic station XIRILNeon-100 produced by XIRIL AG, Switzerland. We used a reagents kit “Magno-Sorb AmpliSens” manufactured by FBSI Central Research Institute of Epidemiology of Rospotrebnadzor, Moscow, Russia. The amount of DNA obtained was stable at 80–100 ng.

Genotyping of the UGT1A1 ∗28 (rs8175347) polymorphism was carried out by real-time PCR on a Rotor-GeneQ detection amplifier manufactured by Qiagen, Germany. We used a UGT-RT50 reagent kit manufactured by EntroGen, USA. All reactions were carried out in accordance with the manufacturer's instructions.

Statistical analysis

Statistical processing of research results was carried out using standard statistical functions of spreadsheets MicrosoftExcel2007 (Statistical Package Microsoft Office 97 for Windows, Redmond, USA) and an online calculator for calculating statistical criteria “Medical statistics” (https://medstatistic.ru/calculators.html).

The frequencies of alleles and genotypes of the rs8175347 marker of the UGT1A1 gene were calculated as a fraction of their total number in the sample. Statistical analysis consisted of comparing two samples for one feature. The trait was understood as the presence (absence) of a certain allele or genotype.

Statistical assessment of the differences significance in the frequencies distribution of polymorphic alleles and genotypes between the studied samples was carried out using the χ2 test. To assess the statistical significance of differences in the mean levels of total bilirubin between groups in order to identify associations of the total bilirubin concentration in the blood with the pathology development we used the Student's t-test. A P-value <0.05 was considered statistically significant.

Results

In the control group, the distribution of UGT1A1∗28 genotypes was: 6TA/6TA (normal homozygote) 30 cases (40.54%), 6TA/7TA (heterozygote) 31 cases (41.89%), and 7TA/7TA (pathological homozygote) 13 cases (17.57%). The distribution of genotypes is shown in Figure 1a. The frequencies of the 6TA/7TA alleles were 61.49/38.51% (Figure 1b). In the group of patients with GS, the distribution of genotypes turned out to be fundamentally different. The overwhelming majority, 104 cases (83.87%), were pathological homozygotes 7TA/7TA, 20 cases (16.13%) were heterozygotes 6TA/7TA. No normal 6TA/6TA homozygotes were found. Allele frequencies were 8.06% for normal 6TA and 91.94% for pathological 7TA (Figure 1b).

Figure 1.

Figure 1

Frequency distribution of UGT1A1 ∗28 genotypes and alleles in the control group and the patients with GS group. а, genotype frequency; %. б, alleles frequency, %. The abscissa shows the number of TA repeats in the UGT1A1 gene promoter, the ordinate shows %. Lilac columns correspond to the results obtained by genotyping UGT1A1 ∗28 in the control group, burgundy columns correspond to the results obtained by genotyping UGT1A1 ∗28 in the group of patients with GS. a, P < 0.001; b, P < 0.001; χ2 test.

Significant differences in the genotypes frequency (ч2 = 95.31; P < 0.001) and the UGT1A1∗28 alleles frequency (ч2 = 62.93; P < 0.001) between the patients and control groups were revealed.

The blood bilirubin level of all project participants was measured. In the control group the average value was 11.90 ± 4.70 мmol/L. At the same time the bilirubin level was significantly different in representatives of three different genotypes.

In the normal homozygous genotype carriers, the bilirubin level ranged from 7.11 to 27.2 мmol/L. The mean value was 10.95 ± 4.55 мmol/L. In heterozygotes, the bilirubin level ranged from 6.89 to 26.31 мmol/L. The mean value was 12.66 ± 4.82 мmol/L. Indexes of mean value of bilirubin level in carriers of these genotypes are within the physiological normal range. In pathological homozygotes, 7TA/7TA carriers the bilirubin level ranged from 7.65 to 38.72 мmol/L. The mean value was 28.66 ± 9.26 мmol/L which exceeds the normal value of total bilirubin for adults (21 мmol/L). Thus in the control group of healthy young men, the level of total bilirubin increases with an increase of the amount of additional TA dinucleotides in the UGT1A1 gene promoter. The ratio of the bilirubin level with the UGT1A1 genotypes for the control group is shown in Figure 2a.

Figure 2.

Figure 2

Bilirubin concentration in the blood of men of the studied groups with different genotypes. Box-and-whiskers diagram. а, healthy men; б, patients with GS. The abscissa shows the number of TA repeats in the UGT1A1 gene promoter; the ordinate shows the level of total bilirubin (мmol/L). The dots show the mean values of bilirubin, the horizontal lines are the medians.

Bilirubin level measurement in the patients with GS group gave the following values: for pathological homozygotes 31.90 ± 11.14 мmol/L, for heterozygotes 25.45 ± 3.32 мmol/L (Figure 2b).

As in the control group in the patients group, the total bilirubin level correlates with the amount of additional TA dinucleotides in UGT1A1 gene promoter. The mean bilirubin level value in patients with GS group was 31.07 ± 9.88 мmol/L which significantly exceeds the total bilirubin level in blood plasma defined as physiological normal range.

Figure 3 shows the mean total bilirubin values in the healthy blood donor group and in the GS patients group.

Figure 3.

Figure 3

Mean total bilirubin level in the study groups. The abscissa shows the groups of project participants; the ordinate shows the average level of total bilirubin in the blood (μmol/L). The lilac columns correspond to the results obtained in the control group, the burgundy columns correspond the results obtained in the group of patients with GS. P < 0.1; Student's t-test.

There were no statistically significant differences detected in the total bilirubin mean levels between the patients and control groups (Student's t-test value 1.75; freedom degrees number f = 196; P = 0.081323; P < 0.1).

Discussion

Hereditary component of the GS development is the presence of pathological alleles in the UGT1A1 gene which encodes (UGT)1A1. This enzyme controls the conjugation and clearance of bilirubin.4,10,11,14,15 The types and frequencies of UGT1A1 mutations vary by ethnicity. Unfortunately there are very few data on the population genetic characteristics of UGT1A1 in Russian residents.18,19 Considering the high frequency of UGT1A1 biallelic variations and, first of all, homozygous variant UGT1A1∗28 associated with unconjugated hyperbilirubinemia as a GS risk marker15,20 we conducted this study to test this polymorphism in 124 patients with a clinically confirmed diagnosis of GS and 74 healthy men in the North-West region of Russia to describe the prevalence of the UGT1A1∗28 allele (genetic variant code is rs8175347) in these groups and in order to assess the clinical significance of such genetic testing for people with hyperbilirubinemia. Therefore the genetic testing data were compared with the serum bilirubin level values for each project participant.

At least one pathological allele of the UGT1A1 promoter was found in 168 of 198 people who participated in the study. This almost 72% of the men tested are found out as a carriers of the allelic variant UGT1A1∗28 (7TA).

Both heterozygous genotype (TA)6/7 and normal homozygous genotype (TA)6/6 were presented in the control group with approximately the same frequency (41.89% and 40.54%, respectively). Pathological homozygotes were recorded in 17.57% of healthy men. In patients with GS the heterozygous genotype (TA)6/7 was significantly less common: in 16.13%; and the wild-type UGT1A1variant was not identified at all (0%). But the frequency of homozygous genotype (TA)7/7 occurrence was 83.87%.

The wild-type allele UGT1A1∗1 6TA associated with normal enzyme activity is the most common variant among the world's population. It occurs in approximately 61% of the Caucasian race.5 In our study, the total frequency of 6TA allele in the control group was 61.49% which almost completely coincides with the literature data. Genotypes 6TA/6TA (UGT1A1∗1/∗1) and 6TA/7TA (UGT1A1∗1/∗28) are found in the Caucasians with frequencies of 34% and 55%, respectively.7 We identified a heterozygous genotype (TA)6/7 in 25.76% of all project participants. In the control group, this index was 41.89% which is slightly below the average for the white population. Wild-type (TA)6/6 occurs in 15.15% of the men tested by us, and these cases refer exclusively to the control group where they accounted for 40.54%.

As mentioned previously, the 7TA allele frequency in our study was 71.97% for both groups with a peak prevalence of 91.94% in the GS patients group. Polymorphism UGT1A1∗28 is recorded in the healthy men group in 38.51% which significantly exceeds this indicator for Europeans (26–31%).7,16 First of all, this is due to the high percentage (17.57%) of the homozygous polymorphic genotype UGT1A1 (∗28/∗28) in this group. The homozygous (TA)7 prevalence in European populations in most cases is in the range of 5–10%.7,15,17,21, 22, 23, 24 However, there are some exceptions. Thus the homozygotes (TA)7 prevalence in Spanish Basques is approaching 15% (14.8%), and in Romania this index generally demonstrates abnormally high values of 32.33%.7,21 About 10% of the US population is homozygous for UGT1A1∗28.6,7 Similar results were recorded in Brazil.21 In the African continent, the frequency (TA)7/7 is 17.9%, in Asia it is 2.6%, in Thailand and in China, it is 1%.10 Thus it can be stated that the frequency of the UGT1A1 pathological genotype 7/7 in healthy men, we tested significantly exceeds the same average indicator for both Europeans and the Caucasian race in general. We should note that in most of the aforementioned studies when assessing the frequency of occurrence of the UGT1A1∗28 polymorphism in various populations the PCR method was used as in present study. It excludes the possibility of associating a wide spread of the data obtained with the shortcomings of methodological techniques. In addition, the data obtained indicate a much wider distribution of the (TA)7 allele in the Russian population than was considered earlier18 and point to an increased likelihood of Gilbert's syndrome developing in of Russian North-Western region residents.

The revealed significant differences in the genotypes frequency (χ2 = 95.31; P < 0.001) and the alleles frequency of UGT1A1∗28 (χ2 = 62.93; P < 0.001) (Figure 1) between the control group and the group of patients with GS confirm the important role of UGT1A1∗28 (7TA) allelic variant in the occurrence of this hereditary pathology. It is obvious that genotyping of UGT1A1∗28 is of great clinical importance and should be used as a prognostic factor for the risk of GS developing. The likelihood of GS symptoms manifestation is noted both with a homozygous pathological genotype and with a heterozygous genotype.7,10,24 In our study, 83.87% of patients with clinically confirmed GS had a homozygous (TA)7/7 genotype and 16.13% of patients are carriers in a heterozygous form. We did not reveal a normal homozygous genotype (wild-type) among patients with GS which corresponds to the literature data.23,25

There is no doubt about the unconjugated bilirubin concentration increase in the presence of additional TA insertion into TATA-box in UGT1A1 gene promoter. Moreover in homozygous for UGT1A1∗28 (genotype 7/7) individuals, bilirubin levels were significantly higher than in individuals with 6/6 and 7/6 genotypes.7,21,26,27 Genotypic and allelic comparisons between patients regarding the presence or absence of mild jaundice in our study also showed a relationship between the UGT1A1 gene promoter 7TA mutant allele and hyperbilirubinemia (Figure 2). The absence of the allelic wild-type (TA) 6/6 variant in the group with GS (0%) confirms its positive effect on the bilirubin level.

In the group of healthy blood donors, the mean bilirubin level values in carriers of normal homozygous genotype (TA)6/6 and heterozygous genotype (TA)6/7 are within the normal range (10.95 ± 4.55 and 12.66 ± 4.82 мmol/L, respectively). Carriers of pathological homozygotes (TA)7/7 bilirubin levels ranged over a wide range from 7.65 to 38.72 мmol/L. The mean value was 28.66 ± 9.26 мmol/L which is higher than the normal value of total bilirubin for adults (21 мmol/L) (Figure 2a). In the group of patients with GS, the mean bilirubin level for both heterozygotes (25.45 ± 3.32 мmol/L) and pathological homozygotes (31.90 ± 11.14) significantly exceeded the upper limit of the norm (Figure 2b).

Despite the significant difference in mean bilirubin concentrations in the two groups participating in the project 11.90 ± 4.70 мmol/L for healthy men and 31.07 ± 9.88 мmol/L for patients with GS (Figure 3) and explicit dependence between polymorphism UGT1A1∗28 and hyperbilirubinemia, we did not find statistically significant differences in mean levels of total bilirubin between the patients and control groups (P < 0.1; Student's t-test).

Apparently only external manifestations (jaundice, fatigue, sleep disturbances, nausea, belching, and so on) and elevated bilirubin levels in the blood with normal liver function do not allow the diagnosis of GS unambiguously. The value of standard prognostic indicators which are calculated on the basis of total bilirubin levels may be exaggerated. Further research is needed to accurately assess this potential source of bias in clinical practice particularly in GS diagnosis. Therefore genetic testing of UGT1A1 should be the most important component of the mandatory tests list when making a diagnosis of GS because it is the UGT1A1 variability that plays a leading role in hereditary hyperbilirubinemia. This primarily refers to the UGT1A1 ∗28 polymorphism however the identification of additional mutations in the UGT1A1 gene may contribute to the correct diagnosis.

Until now, there is no unambiguous answer to the question of whether exclusively UGT1A1 genetic defects are involved in the GS pathogenesis or whether additional factors are involved in this process. For example, for several reasons, red blood cells can breakdown too easily releasing excess bilirubin that the defective enzyme UGT-1A cannot cope with. In addition, the transport of bilirubin to the liver where it is glucuronidated may be impaired. Therefore assessing the UGT1A1 status alone may not be enough to determine the genetic causes of hereditary hyperbilirubinemia in GS diagnosis. In some cases, genotyping of additional genes will be required.

An important reason for the need for UGT1A1 genetic testing is the GS multifactorial nature.6 In particular, the UGT1A1 polymorphisms can be correlated with chronic liver diseases.4,21,28,29 The GS-associated variant of UGT1A1∗28 promoter is a risk factor for the gallstone disease development.28 The incidence of polymorphism (TA)7/7 is significantly higher (P < 0.001) among patients with chronic hepatitis C.21

The data appeared of a number of cancers associated with GS and UGT1A1 genotype evidence.12,30, 31, 32 It has been reported that GS is associated with an increased breast cancer risk developing.30,31 Polymorphism UGT1A1∗28 is a hereditary risk factor for the colorectal cancer development.32 Apparently for UGT1A1 gene pathological allele (TA)7 carriers, the predisposition to oncogenesis is a consequence of a decrease in the carcinogens detoxification12,21 and altered of estrogen levels.33

In particular, the TATAA box polymorphism of the UGT1A1 is a risk factor for benz(a)pyrene (BaP) metabolite-induced carcinogenesis.12

Another example of the UGT1A1 ∗28 polymorphism negative impact is an increased risk of bronchopulmonary dysplasia developing and an increased mortality of premature infants from respiratory diseases.11 An interesting feature of the bilirubin global biological function is its protective effect against the cardiovascular diseases (CVDs) development due to its antioxidant, anti-inflammatory, vasodilating, antiapoptotic, and antiproliferative functions.3,34 Indeed, in people with the UGT1A1∗28 polymorphism, the CVD and coronary heart disease (CHD) risk is reduced by about three times3 and CHD prevalence in patients with GS is 2% compared with 12.1% in the general population.35

The need for a correct and timely diagnosis of GS is indicated by numerous reports linking GS with increased toxicity of some drugs. Although GS is not considered as a dangerous pathology the mutant UGT1A1 genotype can alter drug metabolism reducing the ability to conjugate. That is in patients with GS in some cases, pharmacogenetic risks increase sharply.36 In particular, the UGT1A1 ∗28 polymorphism increases the likelihood of adverse side reactions when using a number of antineoplastic agents.37 Chemotherapy of malignant neoplasms using irinotecan, sorafenib,20,21,38 and belinostat39 is accompanied by a significant increase in the toxicity of these drugs in patients with GS primarily increasing unconjugated hyperbilirubinemia. That often leads to premature termination of treatment.20,21,40 In addition, patients with UGT1A1 ∗28 treated with irinotecan have an increased risk of diarrhea and neutropenia.41,42 It is obvious that empirical dose adjustment is necessary when using chemotherapy for patients with GS.

Some polymorphisms of the UGT1A1 gene worsen the prognosis in the treatment of viral hepatitis C.21 Hyperbilirubinemia has been reported in patients with GS treated with nilotinib for chronic myeloid leukemia.43 Slow excretion of organic compounds such as menthol and indocyanine green also has been reported in patients with GS.21

Pharmacogenetic assays link mutant UGT1A1 genotype to toxicity of antiretroviral therapy containing protease inhibitors atazanavir and indinavir.5,40,44 These drugs induce hyperbilirubinemia in HIV-infected patients with GS by suppressing the activity of uridine-glucuronosyltransferase 1A1 in the liver by competitive inhibition. More than half of the population is UGT1A1 gene pathological homozygous and heterozygous carriers. All they fall into the risk group for side adverse effects which should be taken into account by clinicians when prescribing specific drugs. The need for UGT1A1 gene promoter typing emerges due to the differences in pharmacogenetic effects when using a wide range of drugs: anticholinergic drugs, hormones, contraceptives, cytotoxins, and so on. Research aimed at understanding the interaction between GS genetic predisposition and drug toxicity will help identify individuals with high risk of jaundice developing and will facilitate the treatment regimens correct adjustment.

Summing up, we can draw the following conclusions:

  • 1.

    Polymorphism UGT1A1∗28 occurs in the population of healthy men in the North-West region of Russia more often than in Europeans and the frequency of the UGT1A1 7/7 pathological genotype is significantly higher than the same mean index for both Europeans and the Caucasians in general.

  • 2.

    The revealed significant differences in the frequency of genotypes (P < 0.001) and the frequency of alleles UGT1A1∗28 (P < 0.001) between the control group and the group of patients with GS confirm the important role of the allelic variant UGT1A1∗28 (7TA) in the occurrence of this hereditary pathology.

  • 3.

    Taken together, these data indicate an increased likelihood of developing Gilbert's syndrome in residents of the Northwest region of Russia.

  • 4.

    Genotypic and allelic comparison showed that the level of serum bilirubin increases with an increase in the amount of additional TA dinucleotides in the UGT1A1 gene promoter. However, we did not find statistically significant differences in the mean levels of total bilirubin between the group of patients with GS and the control group (P < 0.1). Apparently, only external manifestations (jaundice, fatigue, sleep disturbances, nausea, belching, and so on) and elevated levels of bilirubin in the blood with normal liver function do not allow for an unambiguous diagnosis of GS.

  • 5.

    Genotyping of UGT1A1∗28 is of great clinical importance and should be used as a predictor of GS developing risk.

Credit authorship contribution statement

I.A.V. conceived the idea for the project, coordinated the study, and conducted most of the experiments. S.E.V. contributed to the preparation of the figures, analyzing the literature data, and writing the article.

All authors analyzed the results and approved the final version of the manuscript.

Conflicts of interest

The authors have none to declare.

Funding

No grant support or any kind of financial assistance from other institute.

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