Objective To explore the effects of bilirubin on myeloid differentiation factor 88 (MyD88)and interleukin-1 receptor associated kinase-4(IRAK-4).Methods Seven-day-old Sprague Daw-ley rats (clean grade),male or female,weighing 12.0 to 15.0 g,were randomly assigned to 6 groups.There were normal saline group(Ⅰ),lipopolysaccharide(LPS)control group (LPS,Ⅱ),15 mg /kg bilirubin con-trol (free-LPS)group (Ⅲ),15 mg /kg group (Ⅳa),30 mg /kg group (Ⅳb)and 50 mg /kg group (Ⅳc), and then subsequently divided into 2 h,5 h and 24 h subgroups in each groups.Some of the 200 newborn rats died amid the experiment.Finally a total of 144 were involved in the analysis of results,and 8 rats in each subgroups.Newborn Sprague Dawley rats were administered at various doses of bilirubin (15 mg /kg, 30 mg /kg and 50 mg /kg respectively)intravenously;1 h after injection,the rats were administered LPS intrap-eritoneally at a dose of 1 mg /kg;MyD88 and IRAK-4 were detected by immunohistochemistry at 2 h,5 h and 24 h after the injection of bilirubin.Results (1 )LPS could stimulate the expression of MyD88 and IRAK-4 in spleen cells (qMyD88 2 h =49.89,qMyD88 5 h =139.54,qIRAK-4 2 h =7.93,qIRAK-4 5 h =24.30,qIRAK-4 24 h =6.97 ,P <0.01 ).(2)Low concentration of bilirubin could promote the expression of MyD88 and inhibit the ex-pression of IRAK-4 (qMyD88 2 h =0.76,qMyD88 5 h =5.05,qIRAK-4 2 h =6.43,qIRAK-4 5 h =22.37,qIRAK-4 24 h =1.50, P <0.01 ).(3)LPS stimulation MyD88 affected by low concentration of bilirubin was not obvious (q2 h =1.48,q5 h =1.45,q24 h =0.10,P >0.05).Effects of medium and high concentration of bilirubin on LPS stim-ulation MyD88 were inhibitory(qⅣb 2 h =42.87,qⅣc 2 h =51.38,qⅣb 5 h =103.61 ,qⅣc 5 h =1 15.44,qⅣb 24 h =1.18,qⅣc 24 h =1 1.66,P <0.01 ).(4)Effects of low,medium and high concentration of bilirubin on LPS stimulation IRAK-4 were inhibitory(qⅣa 2 h =9.52,qⅣb 2 h =14.39,qⅣc 2 h =25.55,qⅣa 5 h =38.83,qⅣb 5 h =54.62,qⅣc 5 h =60.51 ,qⅣa 24 h =2.41 ,qⅣb 24 h =1.47,qⅣc 24 h =7.61 ,P <0.01 ).(5)The inhibition of biliru-bin to MyD88 and IRAK-4 was observed at 2 h,strengthened at 5 h,disappeared at 24 h in low-mid concen-trations of bilirubin(P <0.01 )while still visible at 24 h in high concentration of bilirubin.(6)There was neg-atively correlation between the expression level of MyD88,IRAK-4 and bilirubin concentration(rsMyD88 2 h =-0.86, rsMyD88 5 h =-0.92,rsMyD88 24 h =-0.53,rsIRAK-4 2 h =-0.82,rsIRAK-4 5 h =-0.86,rsIRAK-4 24 h =-0.57,P <0.01).(7) Under the effect of bilirubin and LPS,there were positively correlation between the expression levels of MyD88 and IRAK-4 of spleen cells(r2 h =0.77,r5 h =0.9,r24 h =0.67,P <0.01).Conclusion Bilirubin could inhibit the expression of MyD88 and IRAK-4.As the concentration of bilirubin increasing,its inhibition is more obvious and prolonged.The mechanism that bilirubin affects immune function of newborn rat may be related to regulation of expression of MyD88 and IRAK-4 at toll-like receptor 4 signal pathway.

Objective To explore the effects of bilirubin on myeloid differentiation factor phospho-p38 mitogen-activated protein kinase (p-p38MAPK) and apoptosis in splenocytes of neonatal rats.Methods Seven-day-old Sprague Dawley rats (clean grade),male or female,weighting 12.0-15.0 g,were randomly assigned to 6 groups.There were blank control group (Ⅰ),lipopolysaccharide (LPS) control group (Ⅱ),15 mg/kg bilirubin control (free-LPS) group (Ⅲ),15 mg/kg group (Ⅳa),30 mg/kg group (Ⅳb) and 50 mg/kg group (Ⅳc),and then subsequently divided into 2 h,5 h and 24 h subgroups in each groups.Some of the 200 newborn rats died amid the experiment,tinally,a total of 144 cases were involved in the analysis of results,and 8 rats in each subgroups.Newborn Sprague Dawley rats were administered at various doses of bilirubin (15 mg/kg,30 mg/kg and 50 mg/kg,respectively) intravenously; 1 h after injection,the rats were administered LPS intraperitoneally at a dose of 1 mg/kg;p-p38MAPK were detected by immunohistochemistry;Apoptosis in splenocytes was detected by terminal deoxynucleotidyl transferase mediated dUTP nick end labeling methods at 2 h 5 h and 24 h after the injection of bilirubin.Results 1.Expression of p-p38MAPK in each group:bilirubin in low-mid concentrations of range inhibited LPS-induced p38MAPK activation (qⅣa =20.93,10.37,respectively at 2 h,and 5 h,all P ＜ 0.01 ;qⅣ b =79.97,14.79,all P ＜ 0.01).The inhibition strengthened with increasing concentration of bilirubin.The effect was observed at 2 h,strengthened at 5 h,disappeared at 24 h.Bilirubin in the high concentrations of range stimulated the expression of p-p38MAPK (qⅣc =32.55,19.23,27.72,respectively at 2 h,5 h and 24 h,all P ＜0.01),observed at 5 h,reduced at 24 h.2.Effects of bilirubin on apoptosis in splenocytes:LPS could increased the apoptosis index (AI) of splenocytes(q =54.62,P ＜ 0.01);The AI of splenocytes had no significant change in low concentrations of range of bilirubin (q =43.92,P ＞ 0.05).Low-mid concentration of bilirubin with LPS reduced the AI of splenocytes (q Ⅳ a =4.48,P ＜ 0.01 ;q Ⅳ b =2.07,P ＜ 0.05),while high concentration of bilirubin with LPS increased the AI of splenocytes (q =5.08,P ＜ 0.01).Conclusions Bilirubin in low-mid concentrations of range could inhibit the expression of LPS-induced p38MAPK,while bilirubin in high concentrations of range stimulated the expression.As the concentration of bilirubin elevated,its inhibition was prolonged.Bilirubin in high concentrations of range bilirubin could induce apoptosis in splenocytes.The immune dysfunction in neonatal hyperbilirubinemia may have something to do with the regulation of phosphorylation of p38MAPK and activation of apoptotic pathways.

Neonatal hyperbilirubinaemia, clinically presenting as jaundice, is a ubiquitous and commonly a benign metabolic condition in newborn infants.It is a leading cause of hospitalization of neonates in the first week of life.Serum bilirubin has been considered as the most potent superoxide with the peroxyl radical scavenger activity.However, uncontrolled hyperbilirubinaemia or rapidly rising bilirubin can reach a neurotoxic concentration, potentially leading to central nervous system sequelae.Thus, the health status of jaundiced newborn infants is dependent on striking an appropriate balance between the protective effects of serum bilirubin and the risk of bilirubin neurotoxicity.In order to standardize the management of neonatal hyperbilirubinemia (jaundice), many countries have developed clinical practice management guidelines.This review sorted out and briefly interpreted the main contents of clinical management guidelines for neonatal hyperbilirubinemia drafted by the American Academy of Pediatrics and other countries, aiming to provide references of clinical diagnosis and treatment practice to domestic pediatrician.