Proteinuria in Children Living with HIV on Highly Active Antiretroviral Therapy (Haart)

Objective: To determine the prevalence of proteinuria in children living with HIV (CLHIV) and identify associated factors. Patients and methods: This was a cross-sectional, descriptive and analytical study carried out from April to August 2017 in the HIV care centres in Brazzaville and Pointe-Noire. The study included CLHIV with dipstick urinalysis test “Combur 10 Test ® M”. Results: Thirty seven CLHIV on HAART presented a proteinuria, 21.8%. Children were male gender in 21 cases (56.8%) and female gender in 16 cases (43.2%). Mean age was 10.9 ± 3.9 years. The children were infected with type 1 virus in 35 cases (94.6%), vertical transmission in all the cases (100%). Children were living with HIV for about 2 to 4 years of average n = 18 (48.6%) and they were WHO clinical stage 2 in 18 cases (48.6%). 13 children (35%) had CD4 level < 200 cells/mm 3 . All CLHIV (100%) were on HAART thus 20 (37.7%) on the combination of zidovudine (AZT), lamivudine (3TC) and nevirapine (NVP). This combination AZT, 3TC, NVP was a protective factor regarding the occurrence of proteinuria (OR: 0.43; IC (95%). Conclusion: Proteinuria is less observed in CLHIV on HAART. Systematic screening and early management of proteinuria during follow-up of these children improve their survival.


Introduction
Proteinuria is a renal complication frequently observed in children, including those infected with the acquired immunodeficiency virus (HIV/AIDS) [1]. Renal dysfunction is promoted by viral infection (HIV nephropathy), antiretroviral treatment (nephrotoxicity), but also by infectious complications and associated treatment [1] [2]. Immune activation by HIV leads to a state of chronic inflammation of the renal parenchyma, which may be one of the determinants of proteinuria and microalbuminuria [1] [3]. Diagnosis of renal impairment is based on renal biopsy; otherwise, proteinuria is used as a marker of renal dysfunction. Its occurrence during the course of viral infection is a poor prognostic factor in terms of survival [4] [5]. The prevalence of proteinuria in children living with HIV (PLHIV) has already been reported in the USA [6] and in some African countries [7] [8] [9]. The aim of this work was to determine the prevalence of proteinuria among HIV-positive children in Congo, to describe the epidemiological characteristics of CLHIV with proteinuria and to identify associated factors with the occurrence of proteinuria.

Patients and Methods
We carried out a descriptive cross-sectional study between April and August 2017, in the paediatric care centres of CLHIV in Brazzaville and Pointe Noire. As well as in the biomedical center IRIS for the quantitative assays of proteinuria.
CLHIV aged 2 months to 17 years, with positive HIV serology, regularly followed up, with recent CD4 count (less than 12 months old) were included after informed consent of the parents. Children with renal malformations and those with disease that affects the kidneys were not included in the study. Sampling was simple and consecutive, CLHIV who met the selection criteria were systematically tested on admission with a reactive urine test strip. Thus, 170 children were selected for the study. Data were collected using a pre-determined survey form after consultation of follow-up registers.
The clinical investigation was a three-step process. At the first medical consultation; the interview provided information on the child's civil identity and personal history. The physical examination first permits us to assess the general anthropometric parameters (weight, height and body mass index). Then a complete physical examination to look for opportunistic infections. This was immediately coupled with the urine test strip examination, which consisted of measuring the proteinuria using a "Combur10 Test ® M strip" dipped in a sample of urine freshly collected by the investigator, followed by a reading of the dipstick.
The reading method was visual and colorimetric and was performed by two investigators of the research team who had good visual acuity and no colour vision impairment. The test strip was positive when there was a shift from the initial pale pink colour to purple.
The paraclinical investigation at the second visit consisted primarily of the 24-hour quantitative assays of proteinuria. The test was performed using a "Cli- green. These proteinuria results were then used for further analysis of the associated factors.
The study variables were epidemiological (age, sex, age of discovery of HIV infection, mode of transmission, type of virus, duration of disease course), clinical (the World Health Organization (WHO) clinical stages of the disease) and biological (CD4 cell count, proteinuria quantitative assay. Proteinuria was said to be "positive" when it was ≥1+ on the urine test strip) and therapeutic variables (treatment protocol, duration of antiretroviral treatment as "ART").
Data were entered and processed using SPSS version 20.0 and Microsoft Excel 2010. Qualitative variables were presented in numbers and percentages in parentheses. Quantitative variables were expressed as mean ± standard deviation. The comparison of qualitative variables used Pearson's chi-square test, while the comparison of quantitative variables used Student's t-test. A univariate analysis was used to look for factors associated with proteinuria with calculation of odds ratio (OR) and their 95% confidence interval (CI). For comparisons the significance threshold was <0.05.

Results
One hundred and seventy CLHIV were included in the study, of whom 37 (21.8%) had proteinuria. The sociodemographic, clinical and biological characteristics of all study subjects with an overall renal biological abnormality during this investigation are recorded in Table 1. CLHIV with proteinuria were male in 21 cases (56.8%) and female in 16 cases (43.2%), for a sex ratio of 1.3. Their mean age was 10.9 ± 3.9 years (extremes of 2 and 17 years), with the most common age range being between 10 and 14 years (n = 19; 51.3%). The children were infected with virus type 1 (n = 35; 94.6%) and virus type 2 (n = 2; 5.4%). Transmission was vertical among all HIV-positive individuals, and they had been living with HIV for an average of 2 to 4 years, n = 18 cases (48.6%). The clinical and biological characteristics of CLHIV with proteinuria are shown in Table 2. All children were on highly active antiretroviral therapy (HAART); Figure 1 shows the distribution of CLHIV with proteinuria according to the different types of protocols administered. Factors associated with proteinuria in CLHIV on HAART are listed in Table 3.

Discussion
This cross-sectional study is assessing the prevalence of proteinuria in a cohort of HIV-positive children from Congo under HAART and is also attempting to examine associated factors that may increase the risk of developing proteinuria in these children. The CLHIV management centres selected were those in the Open Journal of Pediatrics two major cities of the country. Congo is a country with a predominantly urban population [10]. The results of this study therefore provide an overview of the problem and can therefore be extrapolated to the national level. The analysis of proteinuria was done according to the standards of the American Society of Infectious Diseases (IDSA) [11]; this made it possible to exclude false proteinuria and thus avoid a bias in interpretation. On the other hand, the failure to perform a renal biopsy in case of proteinuria was a limitation for our study.    The diagnosis of kidney damage is made by renal biopsy; otherwise, proteinuria and microalbuminuria were used. Proteinuria is a sensitive marker that allows confirmation of the diagnosis, referral to a glomerular or tubulo-interstitial cause, and assessment of therapeutic efficacy [5]. The current guidelines for urinalysis are based on the standards of the American Society of Infectious Diseases (IDSA), which recommends regular testing as the stigma of renal impairment, including proteinuria using a semi-quantitative urine test strip and a quantitative 24-hour proteinuria assay [11], due to the lack of specific evaluation criteria for children. This is the approach used in our study, as well as in other African studies [8] [9] [12] where the protocol recommended that a quantitative measurement be performed for all proteinuria greater than or equal to 1+. The  [14]. In fact, the program recommends that initial paraclinical explorations be carried out, including a urine test before starting ART, three months after the start of ART, and then every six months during follow-up, in the absence of alert signs related to renal dysfunction.
The prevalence of proteinuria in CLHIV on HAART in this study was 21.8%.
This prevalence is similar to those reported by Ekulu [12] and others a predominantly female [19]. The mean age of onset of proteinuria was 10.9 ± 3.9 years. In a study conducted in Ouagadougou on the prevalence of proteinuria in children followed for HIV infection, Coulibaly et al. reported a younger mean age of 8.8 years [12].
CLHIV were more frequently infected with the type 1 virus in a proportion of 94.6%, close to that reported by Chaparro et al., 98.6% to [6]. Similarly, Coulibaly et al. [12], Barry et al. in Conakry [20] reported a similar result. These results can be explained by the fact that HIV1 is responsible for the pandemic. We believe that the predominant mother-to-child transmission observed in almost all the children in our series reflects the dysfunctions of the PNLS in Congo, and especially the weaknesses in the strategy for the prevention of mother-to-child transmission of HIV infection (PMTCT). This suggests that up stream screening of women of childbearing age should be stepped up, the PMTCT program should be revitalized, a mechanism should be put in place to trace the children being monitored, and children at risk should be screened early, with a view to the early detection and treatment of kidney dysfunction, thereby reducing the risk of kidney damage. The majority of the children were classified as clinical stage II according to the WHO and had severe immunosuppression with CD4 < 200 cells/mm 3 . This result is superimposable to those reported in the literature. Indeed, the authors state that proteinuria was likely to occur at low CD4 levels [6] [15] [20], and at advanced clinical stages of the disease [6] [15]. Thus, the occurrence of proteinuria appears to be correlated with the severity of viral infection.
The most commonly used regimen in our study combined zidovudine, lamivudine and nevirapine, comparable to that of Coulibaly et al. [12] and Esezobor et al. [7]. The responsibility of HIV infection and the HAART protocol for the occurrence of proteinuria in HIV-infected patients is difficult to establish in this study, despite the fact that ART are deleterious to the kidney, regardless of the duration of disease progression and the time of exposure to these molecules. The protocol combining zidovudine, lamivudine and nevirapine has been the most widely used first-line protocol in Congo since 2010, that is to say at the time of the study.
In this series, we did not find any factors associated with the occurrence of proteinuria. However, the univariate analysis shows that the protocol combining zidovudine, lamivudine and nevirapine seems, without drawing hasty conclu-Open Journal of Pediatrics sions, to be "protective" with regard to the occurrence of proteinuria; consequently, it reduces the risk of renal damage with a positive impact on their survival. The nephroprotective role induced by ART has also been reported by Penda et al. in Douala [9]. But taking into account the small sample size, we suggest that a large scale study should be conducted to better draw conclusions.

Conclusion
Proteinuria is relatively less observed in CLHIV on HAART. Systematic screening and early management of proteinuria during follow-up of this category of children improves their survival. There is no associated factor with the occurrence of proteinuria. However, the protocol combining zidovudine, lamivudine and nevirapine seems nephroprotective.