Melamine

Health surveillance data

Administrative data

Endpoint:

health surveillance data

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

other: Report of a screening study with children. Medical/epidemiological investigation not meeting standards as in guideline studies. The reliability can not be assigned.

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2010

Materials and methods

Study type:

medical monitoring

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Monitoring some parameters of the immune system in children that were intoxicated after consuming tainted milk. Comparison between children with kidney stones and those without them.

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

Melamine

EC Number:

203-615-4

EC Name:

Melamine

Cas Number:

108-78-1

Molecular formula:

C3H6N6

IUPAC Name:

1,3,5-triazine-2,4,6-triamine

Test material form:

solid: particulate/powder

Specific details on test material used for the study:

It is not known if melamine alone or melamine + other compounds, such as cyanuric acid, were consumed by the children.

Method

Type of population:

other: Children having consumed tainted milk formula.

Ethical approval:

confirmed and informed consent free of coercion received

Details on study design:

A questionnaire was designed which included children's general health status, parent's educational level and income, and laboratory test results.

Patients:
We studied 182 children who were screened for urinary stones in West China Second Hospital, Sichuan University, between September 10 and October 31 in 2008. Each of these children had a history of consuming melamine-contaminated powdered formula for more than 30 days. Complete blood count, renal function test, humoral and cellular immune function test, urinalysis and renal ultrasound were carried out. In the end, 12 (6.6%) of these children were excluded from the study because of incomplete data.

Ultrasonography of the urinary tract:
A total of 170 of the 182 children received B-ultrasound with a probe of 3.5-5 megahertz (GE Voluson 730, Acuson Sequoia 512, MYLAB 50-XVision or M5 color doppler) in the ultrasonic department of West China Second Hospital. Ultrasound examinations were carried out by experienced sonographers. The diagnostic criteria was in accordance with that issued by the WHO and MoH of Beijing (Ministry of Health of the People's Republic of China 2008; WHO 2008b). We recorded the results including, location, size, number of stones and hydronephrosis. Children without stones served as the control group.

Laboratory tests
Complete blood count:
Fasting venous blood samples of all subjects were obtained after a 30-min rest in a sitting position. Medical technicians obtained whole venous blood samples using standard techniques. All the subjects' blood samples were analyzed for complete blood.

Immune function tests
Humoral immune function test samples were collected in a Vacutainer tube containing separating gel. These tests included, serum immunoglobulin A (IgA), immunoglobulin M (IgM), immunoglobulin G (IgG), and complement concentration3 (C3) and complement concentration 4 (C4). They were determined on a Hitachi 7600-010 autoanalyzer. Cellular immune function test samples were collected in a Vacutainer tube (EDT A-K2). These tests included Cluster of Differentiation 3 receptors (CD3), Cluster of Differentiation 4 receptors (CD4) and Cluster of Differentiation 8 receptors (CD8) . They were assessed by Becton-Dickinson FACS Calibur.
The character of CBC and blood biochemistry in children changed as the children grew older. According to the reference value of different age groups, the 170 children were divided into three groups: (1) aged < 1-year-old; (2) aged from 1-3 years old; and (3) aged > 3 years.

Urinalysis, microscopy and culture of urine
Freshly voided random urine was analyzed with Gao erbao reagents strips on AJ-4270. Proteinuria was defined as 1+ or more on the dipstick. Every sample was subjected to microscopic examination after centrifugation. Microscopic examination was performed by examining urine on the glass slide and observed 10 visions with high power filed by microscope. In urine, the reference of leukocyte was 0 - 3 per high power field (HPF). Quantitative culture of urine was performed on Columbia blood agar base and on MacConkey Medium using 10 µl standard loops.

Results and discussion

Results:

Population features and ultrasonography results:
There were 96 boys (56.5%) and 74 girls (43.5%). The youngest was 3 months old and the oldest was 5 years old. According to the ultrasonography of the urinary tract of the 170 children, 61 had urinary stones and 109 had no stones. Among children with stones, 38 were outpatients and 23 were inpatients. Most of the children (71.8%, 122 of 170) were aged between 1 and 3 years old. The percentages of children who had stones in each group was 31.6% (12 of 38), 34.4% (42 of 122), and 70% (7 of 10), respectively.

Laboratory results:
Urinalysis results showed that the number of white blood cells in the urine of children with stones was much more than children without stones, especially in the group aged between 1 and 3 years old. None of them had urine protein. Urine microscopy revealed that the white blood cell counts of 16 children (9.4%, 16 of 170) were higher than the upper normal range. Among the 16 children, there were 11 children with urinary stones. In urine sediments of children with stones we found some crystals by light microscopy.
The leukocyte count in peripheral blood had no significant difference between children with stones and children without stones, even though they were divided into three groups. There was also no difference in neutrophilic granulocyte, lymphocytes and monocytes between children with stones and children without stones.
In the humoral immune function test (IgA, IgG, IgM, C3, C4), except for IgM, there was no difference between children with stones and children without stones in each age group. In the group aged < 1 year and the group aged 1-3 years old, serum IgM concentration of children with stones was higher than children without stones.
The concentration of CD3+, CD4+ T cells of children with stones was significantly lower than children without stones, especially in the group aged 1-3 years old. However, the concentration of CD8+ T cell of children with stones was lower for children without stones in the group aged < 1-year-old. In the group aged 3- 5 years old, the concentration of CD3+, CD4+ and CD8+ had no difference between children with stones and children without stones. CD4+/CD8+ lymphocyte phenotyping is the most frequently performed analysis in flow cytometry for monitoring cellular immunology. Thus, we calculated the CD4+/CD8+ ratio between children with stones and children without stones in each group. There was no difference between children with stones and children without stones in each age group.

Clinical manifestations:
Most of the children with stones were managed as outpatients for their non-typical clinical manifestations. They would be administrated for further investigations and management as inpatients if they had symptoms or severe renal ultrasound abnormalities such as kidney stones complicated with signs of obstruction. There were 23 inpatients with symptoms and severe renal ultrasound abnormalities. According to their hospital records, the presence of symptoms included frequent micturition, urodynia, hypourocrinia, fever, and vomiting. Three inpatients were suspected of having urinary tract infection, but their culture of urine results showed that none of them had bacteriuria.

Applicant's summary and conclusion

Executive summary:

Some children who ingested melamine-contaminated powdered formula had leukocyturia, but none had typical symptoms of urinary tract infection. It was assumed that the children's immune systems might have some changes. 170 children were followed up who ingested melamine-contaminated powdered formula. Their blood and urine were investigated, their immune state was observed, and also ultrasonography was performed. In the immune responses of children with stones, immunoglobulin M takes a major immune response and the level of CD3+, CD4+ decreased compared with children without stones. There was no difference in complete blood count between the children with stones and those without stones. It was concluded that leukocyturia had a certain relationship with non-urinary tract infectious renal disease and these children are susceptible to infectious diseases.