Some characteristics of the blood of the guinea pig in experimental scurvy

t'o*u STANFORD UNIVERSITY PUBLICATIONS UNÏVERSITY SERIES MEDICAL SCIENCES Vo¿uun II Nu¡rr¡nn 2 Srudies on By ARTHUR TT. MEYER and LELS M. McCORMICK STÁ,NFORD UNIVERSITY PRESS STANFORD UNIVERSITY, CALIFORNIA tmg CONTEI{TS pAtT ?¡tcE I. Tse Svuprol¡¿tor-ocy AND Gnoss MonpgoLocv or. ExPERTMENIAL Scunvy rN TEE GurrvsA Prc, by Arthu'r W. Meyer.. 7 REFERENCES n P¿erns I axo II, AND E¡rÞLANATToN oF P¡.¿rps. , .. .. .foÜoøíng 44 II. Tss M¡¡¡urp Monpnor-ocv or Exppn¡upnr¡r, Scunvv ¡N TEE Gungæ¡ PrgbyArtlwrl4/.Meyer. ........47 Rornnp¡¡cBs 69 P¡.¡rns III-IX, ¡¡.¡o Exp¡,¡¡v¡.uoN oF P¿¿tBs ..foUcuôtus 70 III. Sor¡n Cs¡n¡,cran¡st¡cs oF TEE Br.ooo oF TED Gurr¡re P¡c ¡¡¡ Exppn¡- ldErvrlr. Scunw, by Lewìs M. McCormôcþ...... 73 Ror¡npncns ......... 106 P¿ern I ¿uo Expr.¿¡¡¿troN oF P¡-¡tn. ....foUauíng tß TL?gI l3 PART III. SOME CFIARACTERISTICS OF THE BLOOD OF THE GUINEA PIG IN Ð(PERIMENTAL SCURVY 1ÀBLE I. II. ÏII. IV. v. VL VII. VIII. IX. X. xI. XII. xilI. XIV. LIST OF TABLES PAGE B¿ooo Cou¡srs oN Non¡rr¿rGu¡rsBe p¡cs... ..... g0 M¡xrl¡u¡¿ ¡¡¡o M¡¡lruuu Br,ooo CouNrs, ETc,, oN Nonu¡rGurr.l¡¡ p¡cs g1 Rsr-¡trow on NuMgrn or L¡ucocyrps ro WBrcnr (on Acn) Su¡¿u¡ny T¡rpw nnoM T¡s¿rs VIII, IX, X, axo XI..,..,.. Su¡rriu¡ny on Rssulrs r¡¡ Snn¡ns 1. . , . . . Suuu¡ny on Crr¡rcas rN Sonr¡s 2..,. Suuuenv M¡on ¡,nou T¡¡¡-¡s V ¿rqo VI. Br-oop Courvrs on Srnrrs I Gurlqr¡ prcs wrr¡r Scunvy B¡,ooo Cou¡vrs or. Spnrss 2 Gurxp¿ prcs w¡r¡r Scunvy. Dar¿y W'a¡c¡ns or Srn¡ns 1.. . . . . D¡rr-y \d'erc¡¡rs o¡. S¡n¡rs 2,..... . .. . . r. suu¡uenv or lNrtrer- ¡wo F¡w¡rws¡c'rs e*o sunv¡v¡ror SBnr¡s 1. . Suuuany o¡. Inrr¡¿¿ ¿¡ro Frw¿¿ Wsrcgrs *wo Sunv¡ver, on Senr¡s 2.. Su¡¿u¡ny or 'Wr¡curs aNo Sunvrv¡ror Srnrrs I ¡No Sen¡as 2 p¡cs. .. 82 83 84 86 88 94 98 102 103 104 105 105 LIST OF FIGURES FIGU* PÂcE I-III. Pr-¡rp I ..... ....foilozaing.l0g IV. Gn¡prrs on w'rcnr ¡wo Rrcrel T¿rupsn¡runn o¡.A¡,r, scon¡uuc AN¡rue¿s 9l V. Gn¡,p¡rs Snowr¡¡c rrrB Brcrxx¡¡lo ¡rqo Ewo on Hrruor,ysrs...... gL vI. Gnep¡¡s srrow¡¡vc cn¡r.rces rN THE Rro-c¡rr, couNr, tr* psncaNr¿cn or H¡uocr-oBrN, AND rns Colon flqoex. gz vlr. Gn¡Bss Rrpnpsewrr¡vc rsB pnncnrqr¡cs or Nuc,ÆerED AND Rnrrcr¡r,¡rm Enyrsnocyrps gz vIII. Gn¡pss B¡spn upo¡v rre Nurrrgen or Lpucocvrps pen cu¡rc Mr¡,r,¡rrnren .A'ND UpoN rsp Nurrspn on Lvrvrpsocvæs ¡¡¡o por,vuoRpEoNUCLEARS rN Tsnrp Huuonno L¡ucocyr¡s...... 93 IX, A RnpnooucrroN o¡. Ar.r. tss GRepns or F¡cunr,s IV-VIII, INc¡,usrlæ.... 93 III. SOME CHARACTERISTICS OF THE BLOOD OF THE GUINEA PIG IN EXPERIMENTAL SCURVY1 By Lnwrs M. McConu¡cr This study of the blood of scorbutic guinea pigs was undertaken at the suggestion and with the co-operation of Dr. Meyer, to determine the presence of hematological changes in a deficiency condition "which, if not true scurvy, is indistinguishable from it at present," according to Holst and Frölich '12. Hess '20 stated that a characteristic secondary anemia accompanies scurvy in man and wrote as follows: "Hemoglobin is diminished far out of proportion to the decrease in the number of red cells. There may be polycythemia. Microscopically, the red blood cells show poikilocytosis, anisocytosis, and a lack of hemoglobin; they are slightly enlarged, with the occasional occurrence of exceptionally large cells resembling the dropsical cells described in connection with chlorosis. Sometimes a few nucleated red cells and myeloblasts are seen. . . . The dropsical cells suggest a disturbance of the salt balance in the plasma. "In some cases we have found a decreased fragility of the red cells, which also has been described in chlorosis. The total number of leucocytes is slightly increased. In our cases the mononuclear cells have averaged 66%. .. . . Labor described an oesinophilia during convalescence, a phenomenon which we have not encountered. Some describe a marked increase in the polynuclear cells, which probably is to be regarded as a reaction to secondary infection. There is indeed a marked difference of opinion in regard to the morphology of the blood in scurvy in adults as well as in infants. . . . . The divergent reports probably should be attributed to the fact that the investigators are describing scurvy in various grades of severity, of different stages of development, or complicated by intercurrent disease." Liotta '23. Í.ound in the guinea pig that the large mononuclear cells increase progressively, that the eosinophiles also increase in a still less degree. However, in some cases in man, Eichhorst '13-'14, and Liotta in the guinea pig also, found a regular diminution of the erythrocytes. Leitner'17 and Montanari t82L regulady obtained leucopenia with some lymphocytosis, in man. These conflicting frndings seem to suggest that the changes in the blood characteristic of scurvy should perhaps be sought in the cell picture rather than in the cell count. l This investigation was supported in part by a grant to Dr. Meyer from the Committee on Research of the American Medical Association. I19eI t73 74 SÎUDIES ON SCURVY Pnocroune My first series consisted of twenty healthy guinea pigs'of mixed strain, of varying age, and of both sexes. These were put on an abundant diet of raw alfalfa, raw barley, and water. Although some vitamine C exists in such feed, we know that animals do not get a sufficient quantity of it while on this diet to prevent relatively early death. A second series of sixteen pigs was divided into four groups of four pigs each, and fed as follows: Series 2, Group l-raw alf.alf.a, water, and orange juice. Group Il-raw alfalf.a, raw barley, water, and wheat germ. Group III-raw alfalfa, raw badey, and water. Group IV-raw alfalf.a, raw barley, water, and greens. Orange juice supplied vitamines A, B, and C to the diet (Sherman and Smith '22) of the first group. Substitution of wheat germ for the orange juice supplied only vitamines A and B (Eddy '2I) to the second group. Groups III and IV were run as controls, the diet of the former being deficient in vitamines B and C and possibly A, as in Series L, and that of the latter being a normal diet containing all of the vitamines. Tests as given below were continued till within several hours before the expected death of an animal, when it was gassed and necropsied, except those in Groups I and IV of Series 2. The procedure used in making these tests \ryas as follows: Both ears of the guinea pig were shaved and washed thoroughly with soap and water before the beginning of the experiment. The washing was repeated as gently and carefully as possible about frfteen minutes before each sample of blood was taken, and this washing was followed by further gentle cleansing with 95 per cent alcohol. A cut was made with a sterile spring lancet and a natural flow of blood obtained. No sample of blood was taken while the animal was excited or struggling, aud all were taken successively and as rapidly as possible for making fragility tests, the estimations of hemoglobin, erythrocyte and leucocyte counts, and smears for differential white cell counts, and for nucleated and reticulated erythrocyte counts. The rectal temperature was taken as the last step. Reo Br-ooo CBr-r. CouNr A certified Leitz hemocytometer was used. The pipette and counting chamber were thoroughly cleansed by a soiution of sulphuric acid and potassium dichromate, water, and alcohol. A sample of blood taken after the first two drops had been removed was drawn to the index "0.5" on the Thoma Blood Diluting Pipette with great care. lMhile doing this, the pipette was held horizontally and at right angles to the line of vision so [200] THE BLOOD /5 that the exact height of the column could be easily seen and the tip submerged to avoid entrance of air-bubbles. If the blood went slightly beyond the mark, the tip of the pipette was touched to a moistened, clean cloth or the tip of the finger until the "0.5" mark was ¡eached. The pipette was then quickly wiped off, and plunged into Hayem's Diluting Solution and this drawn up to the mark 101, with a slight rotation of the pipette, thus diluting the blood t:200. The pipette was shaken vigorously transversely for temporary mixing, after which a heavy rubber band was placed a¡ound the ends and the apparatus was laid down in a horizontal position. After removing the rubber band several minutes later the pipette was shaken transversely for five minutes and three or four drops were expelled and a small drop placed upon the Double Neubauer counting chamber. The counting chamber and cover slip were cleaned and dried so that when they were placed in position Newton's color rings were visible. Immediately after the drops were in place the cover slip was quickly applied in an oblique manner and the Newton color rings again observed. The counting chamber was placed on the stage of the microscope, in a level position, and after about three minutes, to a1lo\4/ for settling of corpuscles, the counting was done provided the corpuscles lryere evenly distributed over the entire field and no air bubbles were present. Otherwise, the pipette was again shaken for five minutes and the process repeated. The counting was made with low-power magnifi cation. The cell count was made in five sets of sixteen squares each, taken from the corners and middle of the counting chamber. The cells on both counting planes rryere enumerated. If the count in the five sets of squares varied more than twenty or the total count in the eighty squares did not vary less than ten, a nerlv sample of the mixture was taken and the whole process repeated. To calculate the number of red-blood cells per cubic millimeter the total number in the eighty squares was divided by eighty to get the average per square. By multiplying this average by 400 (the area of each square being 1/400 sq. mm.) and by 10 (the depth of each square being 1/10 mm.) and by 200 (the extent of the dilution) the number of cells per cubic millimeter was obtained, as indicated in the {ollowing example: 113x4O0x10X200- :.-.-80j:-l- : 5,430,000 According to Gulland and Goodall '25, "'When the dilution is I in 200, the addition of four cyphers to the number of corpuscles counted in eighty squares at once gives the number per cmm." The average difference in a series of preliminary counts made on myself was 2 per cent. lZotl 76 STUDIES ON SCURVY Esrrlr¿.rron or HnMocr-osr¡[ The hemoglobin determination was made with a Sahli-Leitz Nonfade Hemometer. This instrument consists of two solid glass comparison cylinders vertically arranged and a graduated test tube of the same diameter, and a pipette o120 cmm. capacity. The graduated test tube is placed between the standard solid cylinde¡s and the three held in a black frame with a white ground-glass background. A tenth normal solution of hydrochloric acid was placed in the graduated tube to the mark "10." A drop of blood was obtained from the ear of the pig immediately after the specimen for the erythrocyte count was taken and drawn into the pipette to the 20 cmm. mark. The tip of the pipette was wiped off and the contents were blown into the graduated tube. This was followed by three washings of the pipette with distilled water. These washings were mixed with the chamber contents and the whole permitted to stand for two minutes. After this interval distilled water was added drop by drop with thorough mixing, until the color matched the color of the solid comparison cylinders. In this method the hemoglobin is changed to acid hematin, the color of which is light brown and very easily matched. The percentage of hemoglobin is read directly. The average difference in a series of preliminary estimations made on myself was 3 per cent. Cor-on luorx The color index was determined after the usual manner, by dividing the first three figures of the blood count by five and then dividing the percentage hemoglobin by the resulting number. This method is expressed in the following formula: r-f _ %Hb _ %Hb v.¿.--%¡gf-@ This indicates the percentage of hemoglobin contained in each red corpuscle as compared to the normal. \Mnrrr Br.ooo Cnr-r. Couxr The leucocyte count was made with the same hemocytometer used for the erythrocyte count. A sample of blood was taken from the same incision made for the red blood cell count and hemoglobin and the same steps were followed and cautions observed as in the case of drawing the erythrocyte samples. The blood was drawn to the "1" mark on the white-blood pipette and diluted with a 1 per cent solution of glacial acetic acid in distilled water to which 1202 1 T}IE BLOOD 77 enough of methyl green had been added to color the solution a fairly deep green. The acid rendered the nuclear outlines sharp and the methyl green tinted them faintly. The procedure followed was that used in counting the red cells. Cells included in the total twenty-ûve sets of sixteen squares rvere counted. The total number of squares was 400. A mechanical stage was used for counting both the red and white blood cells. By dividing the total number of cells in the 400 squares by 400, the average number of cells per square was obtained. Multiplying this average by 400 (the area of each square being 1/400 mm.) and by 10 (the depth of each square being 1/10 mm.) and by 10 (the extent of the dilution) the number of cells per cubic millimeter was obtained. Example: 70 x 4oqX 10 x 10 : z.ooo per cmm. 400 ''' When the dilution is 1 to 10 the total number of cells in the squares multiplied by 100 gives the number per cubic millimeter. A series of preliminary counts on myself sho,wed an average difference in the counts to be about 5 per cent. Pnrrenerror on Slrcens The slides used for making the smears were thoroughly washed with soap and water, rinsed well, and placed in sulphuric acid and dichromate solution for a couple of days. They were then thoroughly washed with hot water, dried on a clean cloth, and placed in 95 per cent alcohol for about twenty-four hours, wiped and dried over a Bunsen burner and stored carefully for future use. A very small drop of blood was placed on one end of the slide, and half of a white cigarette paper which had been cut transversely was laid down longitudinally and obliquely just behind the drop of blood. As soon as the drop was touched it spread along the under surface of the paper when the latter was gently pulled across the slide. The paper was raised more vertically as it traveled along. In this manner a very even smear was obtained. The smears were fixed in the open air at room temperature for about an hour and placed in acetone-free methyl alcohol. They usually remained in this solution up to one hour after which they were removed and again dried in the open air for a couple of rninutes. They were stained with Jenner-Giernsa stain, a combination of Jenner's and Giemsa's methods. The Jenner solution brought out the neutrophilic and eosinophilic granules, 1203 1 I t.6 3.2 t.9 1.0 142.8 2 5.9 35.0 43.3 I t.0 24.6 3.2 36. f 49.0 t7.0 r.0.6 t8.3 r7.8 12.3 200.0 ?8.5 0 4 Case Number 57 5.4 u.4 3t.4 t0.0 40.L 11.4 50.9 168.1 I10 3.4 2t.8 29.4 t3,6 54.7 146.6 50.8 ?9.6 68 Temperatur€.. .. Erythrocyt€ß.. .. IIemoglobin.... . Color Index,... . leucocytes.. ,. .. Polymorphonuclea¡s. ïrymphocytes, small antl large. ... .. .. ... Eosinophlles.......... Basophiles... . ... ... .. Mononuclears.......,. Transitionals........, &eticulated DrythrocyfÆsi Nueleafed Erythrocytesl., oa o,i E trI' I,oss in Weight. 3.2 25.0 32.0 t.0 86.3 3.8 26,6 34.9 ¡0.0 7.9 3.2 34.3 38.7 7.2 t9.4 28.0 4.0 7,0 3.8 4t.2 48.3 5.ô 25.6 4.0 ts.7 Ër ãl ÊEl ã -õ't cl bt H 51.2 11.7 3f.0 0 50.0 50.0 0 49.8 900.0 200.0 53.3 300.0 69. I 200.0 f00.0 25r.1 166.6 r8.0 58.6 0 75.0 500.0 55.5 57.t 0 114.2 25.0 41.1 0 300.0 t2.2 116.6 54.4 800.0 200.0 44.4 20.o 67.5 74.6 57.t 0 200.0 100.0 s8.4 676.0 78.5 2100.0 0.? 74,4 00.3 33.9 0.4 0.2 2ß.6 25.0 29,4 024.0 19.0 11.3 27.2 0 0.3 0.1 0 0 fsegan. . .. f Comolcte. 4.3 5.0 26.3 12.0 2t.0 24.0 2t,6 3l,3 26.5 23.2 4,3 5.0 4.5 2t.0 9.0 t0.5 8.6 2t.0 4,3 r0.0 t2.0 20.0 9.0 26.3 9.0 2È.2 31.2 n.7 * Percentages calculated from the first and last readings in each "Case." t Here the "difierence" between the first and last readings in percentages is given, the range of error. The erythrocytes of the scorbutics also fell in all cases, the average being 29.9 per cent, while those of the controls showed an average increase of 5.9 per cent. The hemoglobin of scorbutics likewise fell in all cases an average of 38.2 per cent, while that of controls rose an avefage of 1.6 pef cent. The color index of scorbutics was decreased in 96.6 per cent of the cases, it was increased in 1.6 per cent, and showed no change in 1.6 per cent. The average decrease was t2.2 per cent, while that of the controls showed an average decrease of only 3.3 per cent. The leucocytes of scorbutics were decreased in 8.3 per cent o,f the cases and increased in 91.6 per cent, with an average increase of" 79.8 per cent. Those of the controls were decreased in 12.5 per cent of the cases and increased in87.5 per cent, with an average increase of. t5.7 per cent. The polymorphonuclear leucocytes of scorbutics were decreased in 1.6 per cent of the cases and increased in 98.3 per cent, with an average increase of 232.6 pet cent. Those of controls showed an average increase of only 2.4 ,[210) 84 STUDIES ON SCURVY TABLE Suu¡r¿eny or Rpsur.rs rx Se¡.rps 1 Rsconopo rN lBold face frgures indicate o oí OA lo o a€ äÞ o¡ 7o THE BLOOD V Pencnxr¡cps BasBo oH lxrrrerAND FrNAL R¡¡orwcs* decrease; roman, increaseJ Case l¡.umber 11 12 13 11 16 16 fl 18 t9 20 3.8 2.4 85 ò! Þ o ãü 8ð lo u.g !7.8 42.0 25.5 18.0 7.î 49,2 111.0 4.0 4.0 il.3 24.4 46. I3t.7 t8.0 I r.l 4.2 t5.8 5.8 t.0 88.2 3.4 23.1 32.6 t3.4 63.2 4.4 23.0 37.7 17.0 21.1 t.4 2.2 t.0 0 5.4 0.6 29.9 37.3 r0.4 52.9 2.7 23.7 28.7 ô.1 160.7 3.4 24.8 3t.4 9.1 43.0 0 û 0 6 76 95 6 46 36 70 48 100 60 0 0 100 100 100 90 25 95 35 2Ã. 15 35 0 0 90 95 100 0 0 0 6 0 0 0 ^)40 15 20 0 40 10 6 0 136.9 716.4 69.6 nL.l ß2.3 83.8 1ô?.6 L6¿.4 108.0 b 166.0 24.0 7U.7 100.0 0 57.t 9.0 22.2 5t.4 z¡15.3 200.0 6.2 100.0 8.6 2t.0 ¡15.6 88.8 0 166.6 60.0 65.0 50.0 0 66.6 66.6 72.0 100.0 600.0 0 50.0 73.7 1.6 85.3 500.0 2m.0 66.ô 0 60.9 t6.2 0 2û.0 60.0 32,8 69.3 0 67.r 41t.0 83.0 t9. t 100.0 0 0 0 72.0 29.0 0 0 776.0 33.3 1L4.7 n.4 90.3 47.6 81.8 160.7 38.8 72.1 1.1 0.6 10.7 0.1 17.6 0 9.0 31.5 16.4 0.1 8.0 0.6 0 0 0 3t.6 t3.6 26.3 9.0 26.3 15.9 0.2 1.õ 01.5 0.1 9.5 16.ô t8.3 0 15.7 12.0 2t.0 4.7 I l.l 7.6 t7.8 29.4 t9.3 28.2 45.7 38.4 2t.8 30.2 8.1 26.8 0 per cent. The lymphocytes (small and large) of the scorbutic pigs \¡¡ere decreased in 98.3 per cent of the cases and increased in 1.6 per cent, \r/ith an average decrease of.63.7 per cent. The eosinophiles of scorbutics were decreased in 31.1 per cent of the cases and increased in 51.1 per cent. They showed no change in 17.1 per cent of the cases, u¡ith an average increase oÍ.37.6 per cent. In the controls they suffered a decrease in62.5 per cent of the cases, an increase in37.5 per cent, with an average increase oÍ. 12.9 per cent. The basophiles of the scorbutics showed a decrease in 27.7 per cent of the cases, an increase in 33.8 per cent, and no change in 38.3 per cent, \4/ith an average increase oÍ. 44.6 per cent. Those of the controls showed an average decrease of 0.8 per cent. The monocytes of scorbutics showed a decrease in 32.7 per cent of the cases and an increase in 51.1 per cent. There ìMas no change in 16.1 per cent of the cases. The average increase was 18.3 per cent' fn controls there was a decrease in 62.5 per cent of the cases and an increase in 37.5 per cent, with an average decrease of 16.8 per cent. The transitionals of scorbutics decreased in 42'2 per cent of the cases, lzttJ 86 lDemDerature. ErytÞrocyt€s. Eemoglobin.. Oolor Dldex.. Leucocytes... Reticulatetl Erythrocyt€sl. . Nucleated Erythrocytest. . . STUDIES ON SCURVY TABLE Suuu¿xv oF C¡IANcES rN SERTES 2 rr.r P¡ncnr¡ [Bold face figures indicate Group I O¡se Number 7.1 10.0 60.0 200.0 64.6 6¿.5 0.4 0 0 0 9.8 1.0 3.1 2.0 0 0.4 16.0 3.9 33.S 100.0 2r.2 160.0 0.t 0 0 0 12.2 0.6 2.Ê t.9 1.0 1.6 0 2.8 2.2 2.0 6.8 Av ..3 .2 .0 .t2 2.9 0.2 2.5 2.0 0 8.0 t6.2 0,2 0 0 D.D 15.8 @ ot !t ,3Ë I ã ã'l :5() I Él !r 'ã ËB ã9 .bY o! E' ñH Polymorphoûucloars,. ,.. ... .. ...... Lymphocyt4s, smau antl lalgo,.... DosiDophi¡es.... Basophlles..... . l{ononuclearg. . . Transitlonals, . . 28.S 11.4 80.0 40.0 d0¡ | 16.6 8.1 60.0 6ô.6 ít.4 14.2 1.0 6.1 t4. I l.õ t5.0 n.1 0.8 0.2 0 fBegan..., lOonolete 0 00 1.3 Change in welght. 65.õ 26.4 * Percentages calculated from the first and last readings in each .,Case," I This animal was killed for study of early pathologlcal changes not concerned with the experiment. f Here the "difierence" between the first and last readings in percentages is given. increased in 40.0 per cent, and showed no change ìn 17,7 per cent. The average increase was 77.0 per cent. Those of the controls lvere decreased in 50 per cent of the cases and increased in 50 per cent, with an average increase af 4I.7 per cent. The reticulated erythrocytes of scorbutics showed an increase in 100 per cent of the cases, \lrith an average increase of 13.0 per cent. Those of controls suffered an average decrease of 0.5 per cent. The'nucleated erythrocytes of scorbutics \4/ere increased in 78.3 per cent of the cases, showed no change in 21.6 per cent, \4¡ith an average increase of 0.3 per cent. Those of controls showed no change in all cases. Hemolysis began later in the scorbutics in 88.3 per cent of the cases. There was no change in 11.6 per cent. The average decrease was 8.4 per cent. In the controls the average increase i4'as 0.5 per cent. The completion of hemolysis was hastened in 81.6 per cent of the scorbutic animals. No change was established in 18.3 per cent of the cases, and the average del2t2r 12II THE BLOOD VI TÂcEs BASED oN INrrrAL ÂND FrNAL Rp¡orwcs* decrease; roman, increasel Glroup UGroup III Case Nu.mber AV b7 !.2 3.8 x34.4 ¡08.5 . 8.8 tz.5 t7.0 91.9 42.2 21.3 66.6 168.4 35.4 25.0 r00.0 0 t4.2 12.2 2.4 30.0 0 8,8 66.6 6.0 11.6 r0,0 t00,0 30.7 14.2 2.0 7.6 260.0 200.0 73iL6 26.0 20.0 4.7 50.0 100.0 40.0 300.0 87 AY 6.9 .7 40.0 0 t8.7 61.0 .8 0 Group IY Case Number 6 8 40.8 40.0 49.5 /ß.0 t4.0 r5.3 293.9 136.2 4ÍÌ4.1 1ür.0 267.6 48.3 .a 2Íi.8 050.0 3r.3 4{.0 {0.0 49.5 r5.3 9.4 124.1 7U,0 0.4 n.6 8.6 r0.9 30.7 0.4 13,0 4.2 9.0 n.6 o.2 4.4 3.0 4.0 44.0 0.8 7.L 8,0 2.0 72.0 2.24.0 2 0 0.1 0 0 0 86.0 4.7 0 68.4 3.4 20. I 4t.8 25.0 2.4 36.9 38.8 3.0 16.6 695.6 63.9 0 200.0 37.6 0 11.7 6.4 28.5 1.1 0 79,8 33.3 6.3 0.7 .0 .6 10.0 25.0 60.0 20.0 11.4 10,9 .4 11.3 0.2 9,0 tô.6 7.0 8.2 t3.0 20,0 n.0 0.'t 3.6 0,1 9.0 t7.8 n.s t.2 0 0 0 138.3 0.3 0 0.9 9.5 0 3t.0 t0.5 t6.û 0 0 r9. I 26.9 30,0 3r.6 91,7 crease was 14.6 per cent. An average increase of 0.6 per cent was observed in controls. This summary shows very distinct changes in the blood picture in scorbutic pigs, except perhaps in regard to the transitionals, monocytes, basophiles, and eosinophiles. There was a loss in weight in 100 per cent of the scorbutic pigs, the average decrease being 27.8 per cent. (See Table VII.) The controls, on the other hand, all gained weight, the average increase having been 52.8 per cent. Hess '20 stated that a low-grade fever (100-101) frequently accornpanies scurvy in man, but noted that it may not be a true "scorbutic fever" but merely a "condition grafted upon the nutritional disturbance," for it sharply subsides upon the giving of antiscorbutic food. Hess attributed the high temperature to a complicating infection. Whatever the explanation may be, no rise in temperature was noted at any time in these or in any other animals except when antiscorbutic food was given. Hence my results confirm the conclusion of Jackson and Moore '16 that "there is likewise no rise in temperature during the disease" [experimental scurvy] 12ts l AY 9t2 Case Number 10 ut 13 14 b16 9.5 t8.5 26.2 1.3 0 0 0 3r.5 3.2 5.9 9.8 8.0 6L.7 3.0 38.8 5t.9 24.0 68.0 315.2 60.5 60.0 0 0 800.0 281.1 57.9 65.3 0 45.0 42.8 .9 0,2 3.2 33.6 43.3 12.4 71.9 258.7 49.2 13.8 100.0 9.9 I t.4 t0.8 t8.0 23.8 00 00 TABLE VII Su¡¿rr¿¡ny M¡op rnolt T¡sr.Bs V ¡r¡¡ VI, Saowrwc P¡ncn¡¡t¿cs Cs¡Ncps rN TrrE Nonrrl¡¿ Co¡crRor-s ¡r¡o Sconsur¡cs [In columns showing average change bold face ñgures indicate decrease; roman, increase] Scorbutlcsl Percenüage Cases Pereetrtage Câses Percentage Cases Percentâge Cases P€rcentege Cases Percent&ge Cases Average Ctrange 2.4 72.9 0.8 16.8 4t.7 0.6 0.6 62.8 Increase Average Clrange ?.2 29.0 38:2 t2.2 79.8 ?3,2.6 63.7 37.6 44.6 18.3 n.0 18.0 0.3 8.4 r4,6 zt.8 No Obange D€cr€ase Decrease Increase No Change TemÞ€rature.... . Erythrocytes, . ,. , Eemoglobln...,.. Color Index...... LeucocYtes.. ,.. . . o ÈLymphocytes, Eosinophiles.. zí z¿t 87.5 e}.6 0.2 l-r,9 1.6 3.3 76.7 72.6 60 60 62.6 62.6 62.6 60 62.6 76 62.5 12.6 8?.õ 60 õ0 s7.6 tt 31,6 60 % 0 72.6 72.6 lm t2.6 0 0 xt 0 0 0 0 D.6 0 0 0 lm a¡.6 8t,6 100 100 100 96.6 8.3 1.6 98.3 31.1 t7.7 &.7 42.2 8.3 81.6 100 0 0 0 1.6 91.6 98,3 1.6 61.1 33.8 6r..1 40.0 0 0 0 1.6 0 0 0 u.1 u) Fl U H 14 a U z tn c) F N) Àsmaìl Basophiles. Mononuclears. Erythrocytes. Nucleated Erythrocyt€Á... oa .9Þ .ËÞ o! f;å Change it Wetg:ht'.. and large.......... Ð o o 0 76 0 0 0 0 0.5 0100 78.3 0 0 0 0 16.1 17.7 0 21,6 11.6 18.3 fBegan,. .. {Comdete. * Groups I and î Series 1, and 00 IV, Series 2. Groups II and III, Series 2, l.Tor¡lel Oontrols* THE BLOOD 89 in guinea pigs, although a majority of the scorbutic pigs showed a mod_ erate leucocytosis. since this leucocytosis very quickly ends upon the resumption of antiscorbutic feeding, it seems unlikely that it is due to infection, although weakened scorbutic animals well may be more susceptible to it. Gulland and Goodall '25 suggested that "the leucocytes, owing to the hemorrhages, are generally found to be increased, but in the absence of recent hemorrhage . . . . they are diminished." It is apparent from my counts that the polymorphonuclear leucocytes and lymphocytes vary inversely with each other. This is represented in Figure VIII. It should be noted that the only animal (Case { Series 1) with a lymphocytosis had a decrease in polyrnorphonuclears. It is possible that this pig was not normal at the beginning and the conflicting statements in regard to leucocytosis and leucopenia may be due to failure on the part of some investigators to recognize this fact. Changes in leucocyte count are due to changes in the lymphocyte and polymorphonuclear counts to the extent of 90 per cent, the remainder being due to changes in monocytes, eosinophiles, basophiles, and transitionals, but the changes in the eosinophiles, basophiles, monocytes, and transitional ce1ls do not seem to be characteristic. Since the erythrocyte count, the hemoglobin, and the color index decreased consistently, one may conclude that these changes in cell count are due to the deficient diet. (See Figure VI.) The hemoglobin decreased proportionately more than the erythrocytes. (See Table VII.) This agrees with the findings of Eichhorst in man, and of Liotta in guinea pigs. The decrease in red blood cells may reflect either increased blood destruction or decreased production. Since smears of the bone marrow show the presence of an increased number of nucleated reds as compared to normals, it seems that there is an increased blood destruction and stimulation of the marrow, although it may also mean that the erythrocytes enter the blood sooner. The reticulated cells increased 13 per cent in scorbutic pigs, while in the non-scorbutic the average decrease was 0.5 per cent. This increase in reticulated cells may be considered as indicating an entry of younger cells into the circulation. As shown in Figure VII, they began to appear after about three weeks' feeding of the deficient diet. Smears made from the bone marrow failed to show the presence of reticulated cells nor could the very large red cells be found. Blood smears from the same blood, made at the same time, and stained for leucocytes with Jenner and Giemsa and with IJnna's solution for reticulated erythrocytes, revealed large vacuolated red cells, which were found to be the reticulated forms. (See Figures I 'f2tsl 90 SÎUDIES ON SCURVY and III.'¿) Many nucleated erythrocytes could be observed on the bone marrow smears. It would also seem that, whatever the cause of red-cell destruction, the young cells are more susceptible to its influence. This would imply a drop in the erythrocyte count and this is what was found in these experiments. Since I was able to predict the presence of extensive hemorrhages from the red-cell count, or from an estimation of hemoglobin, hemorrhage seems the most likely cause for this destruction. The increase in nucleated reds in the great majority of cases of scorbutics may be due to an increased activity in the bone marro$' as shown by the increase in the nucleated red cells there. As shown in Figures II and VII, nucleated reds appear relatively later in the circulation-about the fourth week of feeding-than do the reticulated cells. My observations on fragility of erythrocytes are in accord with those of other workers (Hess '20) on the same animal. I found a slight decrease in fragility after about two weeks. (See Figure V.) It seems to be a rule that the concentration necessary for hemolysis drops with the hemoglobin and the red-cell count. This is shown in Figures V, VI, IX. In Case 1, Series 1 (Table VIII pp. 94-97) for example, neither the hemoglobin nor ¡ed-cell count showed any appreciable change, nor did the fragility test. The sudden drop of hemoglobin within the cell may leave a correspondingly lower tension. Hence a lower concentration of saline solution witl be necessary to maintain the initial osmotic gradient and to equalize this gradient and so complete hemolysis. The synchronous drop of fragility with hemoglobin suggests the dependence of the former upon the latter. I can confirm the statement of Hess '20 that "the red cells show poikilocytosis and anisocytosis" as illustrated in Figure I. Many cases seem to show a greater tendency toward crenation as the scurvy progresses. Hess stated that "guinea pigs generally die of scurvy after having lost about one-third of their body weight; occasionally the loss is greater, reaching almost 50 per cent." My results are in general agreement with these. The average loss of weight in scorbutic guinea pigs was 27.8 per cent. (See Table XIV p. 105.) I found that loss in weight began about the twenty-second day of feeding (see Figures IV and IX) but the slight increase in weight in the early stages of the experiment is considerably less than would be expected in normal animals. All of these scorbutic guinea pigs died after a period of. 25 to 44 days, an average of 35.9 days. With a very few exceptions the uniformity of duration is surprising (see Table XII, p. 104, and Table XIII, p. 105). This implies a uniformity in the animals, in the conditions, and in freedom from outside influence and infection. 2 Figures I-III appear as Plate I immediately following the text. I2t6 1 lHE BLOOD 91 DAYS ,ô 20 ?5 50 35 40 45 tooo 7æ 09" 630 € i,g ?" s .9 $ s'o : E e tS o sz' b' o A 450 J90 33o Froune,ÎV . Cuives for weight ond recfol temÞercture shown in confînuous ond brckeh lines reopeclively. Frc, IV.-Graphs of weight and rectal temperature of all scorbutic animals. IDAYS .¡6õ .43 .AIt I r-o t5 25 lo J5 40 ¿5 '¿.t .JA a._ .t 5 14 i.42 (, o f6.¿, I d 5 õ o .s6 ä o o o o I o .52 Ao .39 ,tð, FrcuRE, \¿ Continuous ond brckeñ ltni5 ¡ndi@te beginning ord end of hemolyo¡s re6pectively. .æ Frc. V.-Graphs showing the beginning and end of hemolysis at difierent intervals in the experiment. f2t7 1 92 STUDIES ON SCURVY DAYS s,25ô oto s&5 6Ë r.o 5,000 4750i'.-.- :ì 4,500 :¡s c 6 e 65 s d .s 4,2n ¿,mo Þ .86 itr750 Froun¡, VI. 3 500 e_Eryihrcèytes --Henoglobin +.. -Colorlndex J,2S o 3,0oo 4 .?9 F¡c. Vl.-Graphs showing changes in the red-cell count, the percentage of hemoglobin, and the color index. o5 to ¡ã DAYS 20E õos{o r.5 zo d e ù ; .e e û ! g .9 E ú l5 r.of c I d ; e ür s3 o 5 2 r<i 5 FrcuRE vll. - Nuclected Erythro¿yfcs -"' Reticulqted E"rhþcyt6 0 F¡c. VII.-Graphs representing the percetrtage of nucleated and reticulat€d erythrocytes. [218] ÐAYS 200 ãro t5 20 z5 ðO 95 40 -46 2S ^ 2ro 3 a ¡¡ ,*g ¡ ,*å u c c 136'b € É. no? o t s3è F å 60b I 5 z tô I I ¡¿- j ,zo L !É roa z o ¡9 a- '¡ t F¡cuRE VIll. -Leuæcyioe ¡-f,ymphocytes .PolynorFhoMlee I 6 4. Frc. VIIL-Graphs based upon the number of leucocyte. upe. cubic millimeter and upon the number of lymphocytes and polymorphonuclears in three hundred leucocytes. DAYS rôßzo?5 3è FiGURÉ X(. F¡c. IX.-A reproduction of all the graphs of Figures IV-VIII, inclusive. Graph I represents v/eight; 2, temperature; 3, beginning of hemolysis; d completion of hemolysis; 5, erythrocyte count; 6, hemoglobin; 7, color index; 8, nucleated erythrocytes; 9, reticulated erythrocytes; l0, lymphocytes (small and large) ; 11, polymorphonuclears ; and 12, total leucocyte count per cubic millimeter. IldtgsgË {È oFÈ6 rã ts rd : gÊrs$$g {{{{ôô6 À,1.\¿"ü.tú 9È øNq CQÈEAF@ 6A6ôsÈÀ oooôqoô FÉH gÉH BOOOAÉO O@HOBôA e9qooqq @tsts¡¡þ¡ 8r888ä8 tszz) ooa s F o Ë o o o oc a X Oose Sex Dats Tlee SFFEpEp EEEEEEE EFF.FFSF @o@Þþþ@oaa¡Þñti9 Temperature I ÞEeeËËÞ ËÞÞËËËÞ ËËEËËËË ' ãBEggEg ËgEãggg g-g-gE-g-s'Ë Erytb¡ocytes tr o o Þ c) c z ¡t Ø 9'¡ r¡Þ Ëtr t4 \'¡ øH ¡r¡ X I t¡J I co HO zè Fl s. Þs ÈdF o Ø I H Èì U) w ¡( s{eEsEE ssEäëËä seaËËËË Eemoglobin HF ts ÉÉB ÉÉ F dgôô-Sgô hbbbþiih þbbESEb Color l]lalex CËESce"Ë ËcË"ËÞe"ä "ã"È"ãËËËê Leucocytes €esssea Säsepee €ä¡eEas PoIymoU)honuclea!s HÉNþNÑÑ HÉHÉÉgÉ FEiSSEE È*{F8Ê$æ däÊ8399 Lymphocytes Få o á Þ Þ ô o Þ eNsposÈ q@€{Èþa 88ßS$àg Eosl¡ophilês oÈ9ooF0@ sooH0oF ooooQHa Basophlles ËoÉrots5 ËçË6qËË ËFËË668 Mono¡uelears oeqÈow{ oqoo*qË ËËocopq lfranslf,lonals ñÉ Xä*iFi3 äir3;3i iää[3[5òr Reticulat¿d Erytbrocyües îSoooeo eooaoo€, ooooêacñ Nucleat€al Erythrocytes bbbbbbb bb'ÈÈbbh bbbbnbbs Began È1 Þ !e bþþåfebs isþÊ!ßbbb þËþHsess ComDlots -Ã,^uocs No ssrcoJs 00ï 12. r 13 5,220,000 6,180,000 6,090,000 4,620,000 4,4ã0,000 4,020,000 3,290,000 6-15 6-n 6-28 7-L4 ?-19 6-t1 6-24 6-29 ?-9 7-76 ?'18 7-n T-n 6-77 6-tu1 6-W 74 7-t5 7-18 7-Ð 7-n ç7'l 6-2+ 6-25 7-g 7-75 7-78 7-n 7-?9 6-77 6-. 4 6-æ 74 7-76 7-18 7-20 9.2.0 2.10 2i46 å:30 2.4ã 2i70 1:30 12:40 10:45 10:36 10:46 9:00 9:15 8:45 8?45 2:00 11340 11:20 11:35 9!45 10:00 9:30 9:30 2i60 7i9o 1:20 1:40 10:00 10:s0 11:00 10:60 3:30 2:40 2:45 11:00 11:16 11:60 11:30 98.8 99 98.8 98.8 98.8 98.6 96.4 6,000 6,600 9,700 7,600 7,100 6,300 7,0m 6,400 6,200 5,0@ 6,100 5,500 6,200 6,800 6,100 6,?ñ 6,S00 7,2W 6,400 6,100 6,900 6,500 6,8@ 3,700 a400 5,500 4,tw 6,200 4,900 6,000 6,400 2,90O 2,700 8,300 3,6@ 3,500 4,2æ 8,900 3,700 t.2 0.6 0.8 0.? 1.1 0.9 4.8 t.7 0.9 1.6 1.3 1.6 L.2 0,9 1.8 4.0 3.0 2.6 2.7 3.1 2.6 2.8 3.6 8.6 2.7 3.1 2.8 1.9 2.1 7.1 1.1 1.9 1.3 1.4 1.1 0.9 7.2 0.8 .44 .44 .4 .4 .44 .42 .40 .& .44 .L4 .& ,42 .44 .44 .4L .44 .44 .44 .44 .42 .4 .4L .44 .42 .42 L' .42 .42 .40 .42 .44 .42 .42 .42 .42 .42 .42 .42 ,44 .N .40 .38 .40 .óð .38 .a .36 .36 .36 .36 ,34 .36 .cô .36 .36 .36 .36 .36 .36 .& .36 .36 .86 .36 .36 .36 .3ô .34 .34 .36 .36 .36 .84 .96 .36 .æ .30 .86 0 o 5 1 2 4 10 8 I 6 6 I 8 10 8 11 I 8 7 10 9 6 6 4 5 0 2 0 1 1 o 10 1 28 n8 9,36 89 193 153 93 169 179 L74 182 L75 1?8 775 165 146 180 716 l'12 767 173 160 163 n5 ttg ut n5 æ0 918 n9 zSL 4 240 tuò M '34 m 2m 2g 67 ß 49 91 I28 188 98 7@ 96 99 106 96 ó 110 7ZL 98 1lJ|2 99 111 108 118 118 4S 61 gt 6 4S 61 63 48 60 63 4a 61 65 Æ 63 72 .99 .99 .99 1.0 1.0 1,0 .96 .gt .gl .96 .97 .98 1.0 .98 .9õ 1.0 1.0 .95 .98 .gl 1,0 .94 .96 1.1 .s7 1.0 .gl .çñ 1.0 .90 1.0 108 tvà 100 98 g7 84 63 gl 98 100 108 99 99 100 100 93 96 98 100 99 100 oo 101 gt gl 98 94 93 96 100 94 95 7V2 99 97 100 gt 102 99 0 1 0 0 0 0 2 z 0 1 0 0 1 1 2 I 0 0 1 a 2 0 0 0 I 0 0 1 0 2 1 0 0 0 2 1 1 0 8 õ 7 I 10 11 12 3 I 7 8 10 11 13 19 I 8 11 t1 6 I I 8 10 14 I 13 11 12 19 Ð I 8 6 D o 7 0 0 0 0 0 0 0.1 7 I 4 6 7 0 0 0 0 0 0 0 0 1 4 I 6 7 Ò 6 I I 8 11 7 6 6 6 3 T 6 3 8 I 4 3 6 M 14. t\) N \ 98.4 w.2 98.8 99 98.6 98.8 99 99.2 99.4 99 98.8 90 98.6 99 98.8 99 4,900,000 5,ü/0,000 5,2t0,000 6,330,000 5,060,000 4,890,000 5,120,000 6,250,000 4,210,W 4,460,000 4,430,000 5,170,000 4,900,000 6,090,000 4,890,0@ 6,200,000 4,680,000 4,690,000 5,150,000 4,800,000 4,?70,000 4,600,000 5,31o,ooo 4,890,000 1.1 1.0 1.1 .gt 1.0 .99 1.0 .98 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 Fl l¡l tÉ r¡ o o U r98.8 99.2 99 98.8 6 99.2 98.6 90 99.2 99 98.8 99.2 98.6 98.8 99 98.8 4,310,000 6,n0,000 4,700,000 5,000,@ 5,260,000 4,620,000 6,170,000 4,870,000 16... . , F O t02 STUDIES ON SCURVY The "graphs" are self-explanatory and give composite curves of all scorbutic animals in this experiment and show the relation of the time factor to the signs described above. The values plotted are taken from Tables VIII, IX, X, and XI, and are averages for ail scorbutic animals (Series 1, Groups II and III of Series 2). TABLE X Derr-y W'nrc¡¡rs on Srnres 1* Date Case Number 1 4 6o,7 I10 11 72 13 14 15 16 77 18 19 w 7m6 4-10.. &-13.. ç74.. ç15.. &-16.. &-17.. 4-19.. ç2ß.. 4-n.. 4-n.. 4=29.. 4-û4.. 4-26.. +-27. . +-... 4-29.. 4-30.. 5-8.. 54 .. 5-5 .. ffi.. í-',t.. 5-a .. 5-9 .. 5-10.. 5'11, . ú-12.. õ-13.. 5-14.. 5-.4.. 860 860 860 860 860 865 845 830 815 860 880 870 870 865 g',t5 870 855 810 764 7 16 695 655 635 615 600 590 t 790 805 810 815 830 825 770 I@ ?90 805 816 820 820 815 810 ?90 740 765 735 105 650 650 615 60û 580 I 860 atò 860 880 890 9@ 880 860 890 895 905 920 910 910 s20 935 915 906 885 850 815 ?80 'tffi n.t 690 6?5 660 r ?80 765 76 780 790 800 800 w5 ?90 790 185 760 790 16 ?90 7& 175 740 760 735 ?0û 6',10 655 626 615 595 585 f 920 885 88õ 886 890 895 885 8?5 846 870 815 875 890 880 875 876 8?0 846 845 830 816 ?90 '/@ n5 ?05 680 665 f 730 745 ?60 765 765 n0 780 78ã 800 780 796 810 810 810 800 ?90 800 7 740 710 680 645 640 bt5 555 515 f 620 620 630 630 @5 645 oóÐ 645 640 640 655 655 655 665 660 660 650 620 590 570 546 52t 50û 500 f ?60 760 165 730 740 760 740 ï50 740 735 74,5 't50 135 ?30 725 64 615 595 580 565 550 6ß 62tt 626 510 5ù5 505 616 52t 520 520 515 520 530 540 545 530 526 530 4ffi 415 435 4r.0 390 s75 345 silt 310 æ5 w + 650 650 660 610 660 670 q5 685 6',t5 686 680 690 695 705 ?00 645 w 620 590 550 620 495 480 445 430 4't5 400 n5 frs 220 235 2\lr 240 5 240 245 0 240 246 4.0 2û 2t5 2ù5 26 185 185t l'15 185 200 200 zlo 2$ 200 n5 275 270 * IMeights in grams. weighing was done as nearry as possibre at the same time of dav. t Lettuce was added to thè diet rrere. weights ðorisiãerè¿ *itrt ñoi.-isã;ãï6'io fuií"i, tgzø. $ Column ends with death of animalIt will be noted that Series 1 ancl füoups Ir and III of series 2 developed scurvy while Groups I and IV, series 2, rcmained, normal in every respect. It is believed that these results show clearly that vitamine B as found in wheat germ, as fed, cannot in itself prevent the ,,apparent" paralysis or other signs of scurvy. The blood picture as well as anatomical and other findings of the affected animals in ,Group II, Series 2, agreed in all respects with those of Group III, Series Z, and. with Series 1. In addition to the typical signs of scurvy noted in the living animal, the autopsy findings also confirmed the presence of this disease. Gross signs of infec1,2281 646 646 650 650 635 6Xi 630 636 645 645 650 645 655 64û 650 650 640 580 660 545 620 50û 490 900 885 885 895 895 900 895 880 895 900 910 924 92lt 910 930 930 910 910 880 840 800 ?80 7',l0 't15 105 I 865 810 805 ?90 785 ?80 805 800 815 815 82¡i 835 830 8f¿5 826 830 82ù 795 7n 755 ?30 705 686 655 6'36 620 610 + u5 860 860 865 860 850 840 830 850 845 855 850 8601 830 &10 ?85 77t 165 't50 ?30 'tç5 680 665 445 I 708 685 gt6 690 690 685 615 675 vlD 6't6 665 665 650 640 640 626 610 516 500 485 I 665 6'.t5 680 6',t6 685 690 ?00 695 ovc 700 705 ?10 720 770 700 695 680 650 M5 610 590 565 550 5, 52$ t ?10 700 7W 710 72.0 n5 Ì16 ?10 w 720 725 ?20 775 7w 175 n5 7 665 625 600 585 555 540 505 500 485 f 696 695 690 î10 695 '170 685 690 695 690 685 685 695 690 720 695 690 700 695 680 660 620 695 565 550 575 490 485 + 185 t?6 180 195 195 200 206 n5 270 zLo 210 2r0 A5 270 2@ 195 195 180 170t 715 1?0 170 165 160 160 160 740 140 135 THE BLOOD TABLE XI D¿,rr,v W'nrcsts o¡' SBnrrs 2* 103 Case Number Date t9a6 6-1û...... . 6-11,...... 6-72....... 6-14.. .. .., 6-15.. .. . .. 6-16....... 6-17....... 6-18....... 6-19....... 6-21... . . .. 6-23....... 6-24... .. , . 6-25....... 6-96....... 6-29.. .,. .. 6-30....... 7-3....... 7+....... 7-g .....,, ?-10....... 7-:12....... 7-13....... 7-74.....,, ?-15....... 7-16....... 7-r7 .... ,. . 7-18....... 7-19....... 7-2t....... 7-24...,... 7-25..... .. 't-26... ... . 7-n....... 7-30....... a_1 8-3....... 8-6....... LO 615 605 605 595 605 600 610 610 600 620 610 610 615 62A 675 615 610 580 610 620 ô30 630 640 630 630 645 640 660 63û 640 645 640 630 63û 650 665 6',15 690 t 660 655 670 655 670 650 645 640 630 660 660 650 650 650 660 660 660 660 660 660 670 690 690 690 680 685 690 695 700 695 '1].5 ?05 690 675 695 720 725 + 305 sl5 3n 926 340 346 340 350 350 810 355 370 370 sr6 390 390 395 410 400 4ú 445 440 460 465 4i10 470 50û 490 500 490 500 490 520 530 555 57t 605 t 4 665 630 665 660 640 650 660 655 660 670 680 680 6?0 650 640 650 680 680 680 ?00 700 710 690 17.5. 't20 11û 705 696 125 730 140 750 750 't6ù 170 + 730 ?80 160 ?80 790 800 800 780 785 ?00 ?90 ?90 'ì70 195 800 800 810 szo 810 820 815 810 795 790 745 14û 720 1tû ô90 btÒ 640 @0 600 590 t 680 660 660 660 680 680 680 6?0 680 690 685 680 700 685 üa 660 685 700 695 690 otu 640 620 590 565 655 530 520 510 490 4ú 1 805 800 195 't90 800 810 800 790 16 710 795 800 810 800 810 810 820 820 820 810 ?90 ?80 185 754 lsa fl.0 690 665 640 630 OUD 580 I 0 250 0 zl0 270 280 280 290 290 310 335 340 325 340 340 340 360 365 360 380 390 400 400 410 425 425 450 44t 460 ß5 425 460 I 257I10 1L 72 13 14 15 16 8?0 840 800 810 æl 840 830 820 830 835 840 840 830 850 830 830 830 820 860 870 865 860 860 850 780 ?60 725 ,126 700 685 670 650 640 620 600 o o Ë ô 325 315 320 320 330 840 360 360 375 390 410 405 420 420 444 430 445 420 460 465 410 475 480 490 490 500 615 505 600 510 510 f 230 250 0 265 285 290 305 305 324 340 360 æ 365 380 400 410 430 450 445 470 460 475 490 515 510 530 515 530 540 550 560 + I + s1 1vì/eights in grams. Weighing was done as nearly as possible at the same time of day. Individual dõse of orange juice (3 cc.) given here. Column ends with death of animal. Orange juice (2 cc.) given by pipette daily from this date. rtreights considered to July 926. Í22e J 430 44t 430 450 460 460 490 414 490 480 510 5r.0 505 510 510 530 520 540 530 545 540 550 550 510 í',t0 580 580 580 ÐtÐ 580 5T5 585 Ì 730 740 7 710 n0 680 ?00 700 690 700 690 700 690 700 n0 775 115 1b 700 tÌ0 't 705 690 680 635 600 570 55ú 550 540 520 500 t 850 840 830 830 835 820 780 790 710 s25 825 830 830 8n 825 820 425 8% 87,0 780 7',10 ,t90 't85 770 740 n6 695 690 665 640 615 595 t 495 505 500 500 52ú 530 530 620 520 52ù 530 540 560 550 55ù 560 565 580 555 550 550 630 490 455 440 +20 465 453 485 410 425 430 420 410 400 380 380 365 340 340 355 360 gt6 386 390 405 395 t04 STUDIES ON SCURVY TABLE XII Surruery o¡ l¡r¡rr¿r, ei.¡o F¡¡r¡¿ Wrrcsrs ¡rvn Sunvrv¡r, on Snn¡¡s 1 C'ase No. Sex tion were absent in all scorbutic guinea pigs, although the possibility was borne in mind. Suryival iD Days 29 34 34 oÕ ?)D 33 ðÕ 29 25 9Ð Ðo 30 m 34 33 31 34 31 2f3 28 24.0 2I.6 31.3 26.5 2ß.2 25.t 29.4 26.2 3L.2 n.7 155 195 n0 2L0 2W 195 255 23t 220 255 2t5 72t 2t5 24t 250 t4ú 2rt 22t 15 39 490 705 590 580 660 585 610 M5 485 665 515 500 545 285 400 õn 485 485 YA 175 64ú 900 860 790 860 780 865 875 705 920 73t 62û 760 225 650 665 695 710 185 2t5 l' r. rl r }' M rl xl rl I' M ¡ F M M r' I. F xl F 29.4 19.3 28.2 4.7 38.4 21.8 30.2 31.6 8.1 18.1 * Time and weisht factors considered to date of addition of lettuce. (See routine weight chart -Series t.) Í2so 1 Co¡¡cr-usrous . 1. Definite cytological changes occurred in the blood of guinea pigs in experimental scurvy. These changes became evident after ien duyr L our scorbutic diet. 2. They consist of a decrease in red blood cells, hemoglobin, and color index, an "appatent" decrease in fragility, a relative decreaie in the number of lymphocytes, an absolute increase in polymorphonuclears, and an increase in reticulated and nucleated red blood cells and leucocytes. 3. The eosinophile, basophile, monocyte, and transitional cell counts were not considered characteristic of scurvy. Weisht at Beginning of DxperlmeDt in Grams lryeight at End of Dxperiment in Grams Weight lost ia Grsms Percentag€ Weight Losü Oase Number THE BLOOD TABLE XIII Sultm¡ny oF INrrrAL AND FrNAL WBrc¡¡rs AND SuRvr\rAror Srnrps 2 Weight aü Beginning of Experiment ln Grams Sex I lPerce.ntI age leain 'Weight Gain in Graüs Weighü at End ot Experiment i¡¡ Grams 105 19elg:bü Lost Percentlà Grams age Lost Duretion h Da,ys Gnoup I IIGnoup III Gnoup IV Gnoup 1. .) 3. 4. 5.... 6.... 7.... 8.... r F xt rr 690 725 605 770' 600 590 365* 500 595 4.6ú 580 585 560 510 460 615 660 305 665 870 730 495 730 850 680 805 430 244 325 240 43 43 42 4L 7õ I t., 65 I 9.8 200 I as.s 105 I rs.s f t..... 1.... I l tn 37 qn óa 36.0 133.3 58.4 91 .7 155 320 190 220 l..lr t...t....t l:::l::::l r' I' r' r' r' F r' I' M ¡ ¡' I I 40 39 M û¡ t]¡ 36 37 31.0 19.1 26.2 31.5 30.0 31.6 27.9 27t 140 130 230 255 215 225 9. 10. 11. t2. 13. t4. 15. 16. + Time and weight factors co¡sidered to date of addition of orange juice to daily rations. (See routine weight chart-Series 2.) TABLE XIV Surrtrrtenv on W¿rcnts ¡¡¡o Sunv¡v¡ror Sunrps 1 ¡.Np Senrrs 2 Prcs 1. I .) 2. .) Series GrouÞ Average Itreighü I,ost ln G¡ams 193.9 792.5 ?ä:l Avsrage I,oss 7o 26.8 26.9 29.8 1231 l Avsrâge Weight Gain in Grams Average Galn Vo 25.8 79.8 Average Duratiot in Days 3t.2 40.0 36.6 42.2 37.0 il m I w2 2 106 STUDIES ON SCURVY REF'trRENCES FOR PART III Bnrne, Mentrw, 7892. "Beiträge zur Kentniss d,er zahl und Massverhältnisse der rothen Blutkörperch en," s ckwalb e's M or þh olo gische Arb eit en, !, ]H.eft 2 : 207. Borr,rNcrn, Hnnrvre¡¡ JonN, 1919. Ma,.ual of Løborøtorg Diagnosis, p, 209. Medical Standard tsook Co., Baltimore. BuRNom, S¡¡¡uBr. H., 1904. "A Study of the Blood of Normal Guinea pigs,', Iournal, of Med,icøl Reseørck, ll: 537. connsrerw, J., uncl zvNrz, N., 1884. "IJntersuchungen über das Blut, den Kreislauf und die Athmung bein Säugethier-Fötus,,, Archia für gesørnte Physiologie,34:173. Eoov, \Mer.rnn H., 1921. Tke vitanoine MønuaL p. 52. williams & wilkins Company, Baitimore. Ercnrronsr, E. L. H., 1973-14. Trøttato di pøtologia e Teraþi.ø sþeci.ale Medicø (Milano, Soc. Ed.), IY,73. Evnr, J., 1899-1900. "The Temperature of the Normal Guinea pig,,, Jowrnøl of Ph,ysiology,25: 24. Goooar,l, Alrxeuorn, 1910. "The Number, proportions, and characters of the Red and \Ä/hite Blood corpuscles," Iowrnø|, of patkol,ogy and, Bøcter,í.ology, 14: 195. Gurr,arvo, Lovrr,r. G., and Goooelr,, Ar,nxeNorn, 1925. Th¿ Blood,, third, edition, pp. 8, 16, 101-102, arrd 289. E. B. Treat & Co., New york City. lfevrrr, Groncns, 1889. Dw sang et ses alterations anøtotnique.s (paris), pp, 769-176. I{rss, A¿rnro F.,1920. Scuray, post ønd, present, pp. 139, l4l, Z0g,2I0, Zll, and 21;6. J. B. Lippincott Company, philadelphia ancl London. Ho¡-sr und Fnölrcn, 1912. "IJeber experimentellen Skorbut. Ein Beitrag zur Lehre von dem Einfluss einer einseitigen Nährung,,' zeitsch,r,ift fi,ir Hygiene wnd, Inf ehtionsþranþheiten (Leipzig), 72 : 1-120. Kunrorr, cited by Ennr.rc¡r u¡ro L¡z¿.nus, 1901. Die Anaernie, Nothnagel, Speciell,e Pøtkologie und, Tkerøþie, VIII, 56. Lnn, Anrnun Bolr-cs, 7924. The Microtonoísf,s vade-Mecwm, eighth edition, p. 186. P. Blakiston's Son & Co., philadelphia. Lerruon, P,, 1917. "Beiträge zur Hematologie und Iíinik des skorbuts,,, LI/iener þ|,,ínische Wochenschr,í.fú (Wien), pp, 928-982. Lrorra, D., 1923. "Experimentai Scurvy. III. Modification in the cell Elements of the Blood," Arck'iaio d,i Farynacologia Sperimentale (Roma), pp.36,26. Lucte, S. P., 1926. "White Blood Cells of the Normal Guinea pig," proceed._ ings of tke Society f or Erþeriøtental Biology and, Medicine, Z4l. No. Z. Lvous, \Mrr. R', and ve¡¡ oE cenn, F. RnlrÉ, 7922. "The Blood of the Normar Guinea Pig," Proceed,ings of the society for Erþeriøcental Biorogy ønd. Med,icíne,25: No.2. 1232 1 TIIE BLOOD ro7 Melessrz, 1872. "De la numeration des globules rouges du sang,', Conoþtes rendus d,e fAcad,érnie des Sciences (Paris), L:75,7528. Moxt.lwenr, lJ., 1821. "Della Formula Ematica Nello Scorbuto,,, Reztistø di Osþed,øIiera (Roma), pp. 407411, Moonn, Josren J., and Jecrsox, Letla, 1916, "Experimental Scurvy Produced in Guinea Pigs by Milk and l\{ilk Products," Journal, of the Am,erican Medical Associøtion, 67 z 7933. PnMav, cited without reference by Evnr, J., 1899-1900. "The Temperature of tlre Normal Guinea Pig," Iournøl' of Physí'ol'ogy, 25: 24. Prrrs, cited without reference by Evnn, J., 1899-1900. "The Temperature of the Normal Guinea Pig," Iournal' of Pkgsiolog!,25: 24. Rrcner, cited without reference by Evnr, J., 1899-1900. "The Temperature of the Normal Guinea Pig," Iournøl' of Physiol'ogy,25: 24. Rrnoen, H,, 1892. Beiträge zur Kentniss d'er Leucocyten und, aerwand,ter Zwstänile d'es Blutes. Leipzig. Srrrnuen, H. C., and Surrn, S. L., 1922. The Vitø*cins, p. 2lI. The Chemical Catalog Company, Inc., One Madison Ave., New York City. Tooo, Jenrns Certrnell, 1924. Cl,inical Diognosis by Laborøtorg Method's, fr'fth edition, pp. 379-380. W. B. Saunders Company, Philadelphia and London. 12331 PLATE I, PART III, AND EXPLANATION OF PLATE Plare I, Panr III F]GURE I. Cell vacuolation, poikilocytosis, arrd arrisocytosis in scurvy II. Reticulated red bloocl cells... III. Nucleated retl bloor1 cells... ?AGE 89 90 90