Pediatr. Res. 14: 1026- 1028 ( 1980) granulopoietic factor(s) neutrophilia neonate

Neonatal Neutrophilia: Possible Role of a Humoral Granulopoietic Factor

Yl

John A. Askin Pediutric Reseurch Lahorato?. Kuplun Ilo.rpito1. Rehovot, Isrcrel

Summary within 12 hr of birth and on the fourth day of life and from 13 of them on the 14th and 28th days as well: Twenty-four-hr urlne During the first days of life, newborn infants have leucocytosis collections and serum samples were also obtained from 10 healthy with marked mrtrophilia and a "shift to the left." the mechanism adults who have served as normal controls. for which is as yet unknown. In an attempt to elucidate whether Urinary and serum CSA levels were determined by modifica- humoral graRvlepietic factor(s) plays a role in this phenomenon. tions of the methods of Robinson ( 10, 11). In brief. 0.1 ml of d serial measurements urinary and serum colony-stimulating ac- dialyzed serum or 0.15 ml of dialyzed sedimented filter-sterilized tivity levels were made in healthy newborn infants and normal urine was placed in triplicate 35-mm plastic Petri dishes (Nun- okler controls. Twenty-four-hr urine collections, serum samples, clon). To this was added a mixture of 0.3% agar in McCoy's 5A adcomplete counts were obtained from 30 full-term normal bled medium containing 10" nucleated cells from the infants 24 hr ad4 days after delivery and in 13 of them on the femurs of C3H/bl mice. The plates were then incubated at 37OC 14th and 28th days of life as well. Specimens were assayed for in a fully humidified atmosphere containing I0"r CO,. Colony their colony stimulating activity levels by their ability to stimulate counts were performed after 7 days as an indicator of the level of bone marrow cells from C3HB mice to grow into colonies in soft granulopoietic activity in the specimen. Only colonies containing agar. Elevated . band form, and counts were 50 or more cells were counted. Previous data have shown a linear found during tbe first day of life, which gradually decreased relationship between the number of such colonies and the amount thereafter. Serum and urinary colony-stimulating activity levels of granulopoietic activity present, allowing quantitation of the were significantly increased (3- to Sfold) over the controls on the CSA present in the serum or urine sample (13). The CSA activity first and fowth days of life, ktdeclined to normal values by the was expressed as the mean number of colonies from triplicate 14th and 28th days. plates per 0.15 ml of 24-hr urine or per 0. I ml of serum (10. 1 I). Speculation RESULTS The rise in ncutrophils and band forms in the newborns is Table I presents the results of total neutrophil. band form, and caused by stimulation of granulogoiesis which is regulated by monocyte counts of the entire study group. In agreement with colony stimulation activity, and which probably occurs during the previous data (8. 14). neutrophilia with "shili to the left" was late stage of pregnancy and the first postnatal days. The possibility d prominent on the first day of life and decreased gradually there- that grodmctiolr these elevated colony stimulation activity levels after. was pronounced during the first 2 wk and is either by the newborn's or the placenta remains to decreased by about 50ri by 4 wk of life. Also, a significant number be determined. of metamyelocytes and a few could be observed in most newborns-on the first day of life.' ~~~k~dneutrophilia is known to occur in newborn infants Figure I shows the mean levels of urinary C'SA on the first. during [he first days oflife, At the same there is a fourth. 14th. and 28th days of life as compared with the levels of increase in the numbrsof band forms, metamyelocytes, and normal adults urinary CSA in our laboratory. At the ages of I monocytes, This initial peak with the exception of monocytes and 4 days. mean urinary CSA levels were 49 f 27 and 32 -t 21 gradually falls the end of the firstwk of life when the colonies per 0. IS ml, respectively, compared to the normal values lymphocyte becomes the predominant cell (8, 14).The mechanism of 95 8 colonies per 0.15 ml (P < 0.001 ). By the 14th and 28th for phenomenon is obscure: overprod- days mean CSA levels decreased almost to normal (I2 f 10 and uction nor cell displacement from storage or marginal pools has I per ml, respectively). been proved to be the cause. In Figure 2, the individual pattern of urinary CSA levels in the last decade, a method for short-term marrow culture in each of the 13 newborns who were lcllowed serially for 4 wk are soft agar has kendeveloped enabled the identificationof presented. Almost uniformly. there were peak C'SA levels in the first an ,,, vi,ro granulopo,et,c factor colony-stimulat,ng 24 hr. decreasing somewhat by the fourth day. and dropping (CSA) in the serum and urine of animals and men (10, 11). "gnificant'y at Ihe age of l4 and 28 H~~~~ CSA can be assayed with mouse bone marrow cells as Figure 3 shows the mean levels of serum C'SA of newborns as target cells (13). [he present study. the levels of urinary and compared with the normal adult serum CSA levels In our labo- serum CSA in full-term newborns were serially assayed to deter- ratory. On the first and fourth days of life. mean serum CSA levels mine [he role of humoral granulopoietic hctorin neonatal were 26 f 16 and 21 & 12 colonies per 0.1 ml, respectively. again neutrophilia. showing a highly signifcant increase in CSA as compared with the normal values of 5 f 4 colonies per 0.1 ml (P < 0.001). On the MATERIALS AND METHODS 14th and 28th days. mean serum C'SA levels dropped to 15 5 I2 colonies per 0.1 ml. still higher than in the controls but the Studies were made on 30 healthy full-term newborns, with difference being statistically insignilicant (P> 0.02). A significant informed parental consent. Complete counts. serum sam- correlation between urinary and serum CSA levels and neutrophil. ples, and 24-hr urine collections were obtained from all newborns band. and monocyte counts was not demonstrated. This is not I I)2h NEONATAL NEUTROPt1ILIA 1027

Table 1 Summary of ah,olute neurrophrl, band-form, und monoci3recounty m [hefirti month of newborn Ife

Age In day\ .-

Neutroph~l\/mm' Band forms/mm'

' Mean + S.D

M Wh 4aMY 14UMY .BthMy NORMAL I1 NEWBORNS CONTROL Fig. I. Urinary C'SA levels in 30 full-term newborns dur~ngthe first 4 DAYS wk of life. i,mean: shuded areas, ?S.D. k~g.2. Ser~alurlnary C'SA determ~nat~onsIn I3 newborns. L~nesindi- vidual patients. unexpected because diurnal fluctuations known to occur in leu- kocyte counts of the newborn could account for this lack of correlaton. the same time, the 4 days duration of the neonatal neutrophil rise appears to be too long to be a release or displacement phenome- non. which regularly takes a matter of hours (I). Theoretically, the postnatal elevation in . band The results of the present study demonstrate a significant ele- forms. and monocytes in the peripheral blood. which were also vation of granulocytic CSA in the urine and serum of normal- found in the present study, could be attributed to one of several term newborns during the first 4 days of life. together with mechanisms. These include overproduction, release increased neutrophil counts. Several lines of evidence. such as of ccllh fruni granulocytic niiirrow rcscrvch <)r frt>nlthc nli~rginiiting CSA injections in mice lc:iding to neutrophilin (7)and the dem- pool, and hemoconcentration. Inasmuch as this neutrophilia in- onstrations in CSA levels corresponding to neutrophil counts in cludes a simultaneous rise in bands and monocytes with a decrease various neutropenias (5). strongly suggest that CSA might be a in the number of lymphocytes (8). it seems unlikely that hemo- true in vivo granulopoietic factor. It is suggested. therefore. that a concentration could be the true explanation. Xanthou (14) favors humoral factor regulating granulocytic proliferation might be the the "displacement" theory rather than overproduction as the most main cause for the production of neutrophilia in the newborns. likely cause because in her studies she found only a small rise in The finding of elevated CSA levels along with a marked neu- neutrophil precursors in newborn blood, however. certain features trophilia during the first wk of life was not unexpected. Apart of neonatal neutrophilia point to an increased granulocyte pro- from previous observation indicating a very active granulopoiesis duction as the most likely mechanism. Most workers in this field in the bone marrow and peripheral blood of the newborn infant have noted the marked shift to the left. increased numbers of during the first wk of life (8). recent studies (6. 9) have demon- metamyelocytes, myelocytes. promyelocytes, and even blasts in strated an increased concentrations of granulocytic colony-form- the peripheral blood during the first postnatal days (8). Bone ing cells in cord blood. indicating an expansion and proliferation marrow aspirates during the first wk of life shows marked cellu- of granulocyte committed stem cells in the newborn. The cell or larity, with increased numbers of granulocytic precursors (12). At cells which are involved in the production of increased CSA in 1028 BARAK ET AL monly found in pregnant women at term (4) and to the neutro- philia of the newborn infants.

REFERENCES AND NOTES

I. Boggs. D. R.: Phys~ologyof neutrophll prol~ferat~on,maturation and clrculatlon. Clin. Haematol.. 4: 535 ( 1975). 2. Burgess. A. W.. Wilson. E. M. A.. and Metcalf. D.. St~mulat~onby human placental condltloned med~umof hemopo~et~ccolony formallon by human marrow cells. Blood. 49: 573 (1977). 3 Chewenlck. P. A,, and Lobugllo. A F : Human blood monwytes: st~mulatorsof granulocyte and mononuclear colony formallon m vrrro. Sc~enceI1 (Wash. D. C.), 178: I64 (1972). 4. Efrat~.P.. Presentey, B., Margal~th.M.. and RoLensLa~n.I.. Leukncytesofnormal pregnant women. Obstet. <;ynecol.. 23: 429 (1964). 5. Greenberg. P. L.. and Schner. S. L.: Granulop~esisin neutropenlc d~sorders. Blood. 41. 753 (1973) 6. Knudtzon. S.. In vrrro growth of granulocyt~ccolon~es from clrculatlng cells in human cord blood. Blood. 43: 357 (1974). 7. Metcalf. D.. and Stanley. E. R.: Haematolog~caleffects on mlce of pan~ally purlfied colony st~mulatlngfactor (CSF) prepared from human urlne Br. I. Haematol.. 31: 481 (1971). 8. Osk~.F. A,. and Nalman. J. L.: Normal blood values in the newborn per~od.In: Hematoloerc-. nroblems In the newborn. nn. . L 30(W. B. Saunders, Ph~ladelph~a. 1972). 9. Prmdull. G.. Prlndull. B.. and Meulen. N. V. D.: Hematopo~etrcstem cells (CFUC) In human cord blood. Acta Paed~atr.Scand., 67 413 (1978). 10. Roblnson. W A.. Bradley. T. R.. and Foster. R.: Bone marrow colony st~mulat~ng Fig. 3. Serum CSA levels tn 30 full-term newborns during the first 4 actlvity In human sera. Br. J. Haematol.. 15: 147 (1968). wk of life. i,mean; shaded areas. % S.D. I I. Roblnson. W. A,, and P~ke.B. L: Leukop~etlcactlvlty In human urlne. N. tngl. J. Med.. 282: 129 1 ( 1970). I2 Shap~ro.L. M.. and Bassen. F. A : Sternal marrow changes durlng the first week of Ilfe. Correlation w~thper~pheral blood findings Am. J. Med. Scl.. 202 341 the newborn are unknown, and this important fact remains to be (1941 ). elucidated. However, it has been shown that monocytes are the 13. Stanley. E. R.. and Metcalf. D.: Partial purification and some propenles of the main source of CSA in man (3). Inasmuch as monocytosis has factor In normal and leukemtc human urlne sttmulatlng mouse bone marrow colony growth m vrrro. Aust. I. Exp. B~ol.Med. Sci.. 47. 467 (1969). been consistently found during the neonatal period (8, 14). the 14. Xanthou. M.: Leukocyte blood plcture In health full-term and premature bab~es possibility that monocytes are the source of the increased CSA during neonatal period. Arch. DIS. Ch~ld..45: 242 (1970). levels must be raised. However, another fascinating finding has 15. Requests for reprlnts should be addressed to: Yigal Barak, M D. Ped~atnc been recently reported, indicating that human placenta (2) is a Research Laboratory. Kaplan Hosp~tal.P. 0. Box I. Rehovot. Israel. 16. Thls research was supported by a grant from the Ch~efSc~ent~st's Off~ce. Ibrael very potent source of granulocytic CSA. It remains to be deter- Mlnrstry of Health. mined by further studies involving pregnant women and their 17. Rece~vedfor publlcat~onAugust 27. 1979. babies whether placental CSA is related to the neutrophilia com- 18. Accepted for publicat~onNovember 13. 1979

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