- Email: [email protected]
Investigation of the representativeness of a single endometrial sample and the use of trichrome staining to aid in the detection of endometrial fibrosis in the mare
THERIOGENOLOGY
INVESTIGATION OF'IHE REPRFSENTATIVENESS OFASlNGLE EN~~ALSAMPLEANDTHEUSEOFTRICHldCMESTAININGTOAID INX-IEDETECI'IONOFENLXMlWRIALFIBROSISINTIiEMARE T.L. Blanchard,1'3 M.C. Garcia:, L.D. Kintner' and R.M. Kenney 1 Hofman Center for ReproductiveStudies New BoltonCenter - Universityof Pennsylvania KennettSquare,PA 91348 and 2veterinaryMedicalDiagnosticLaboratory Universityof Missouri-Colmbia Columbia,MU 65211 Receivedfar publication: JuZy 25, 1986
Accepted: August 5, 1987
The uteri of fivemares were rmovedand er&xnetrialsampleswereprocured fran 12 specificlocationsin the uteri and the saq~leswere processedand duplicatesectionswere stainedwith henatoxylinand eosin (H&E)or Masson's trichrcm stains. The sampleswere interpretedin a blind manner by one person, and pathologicchangeswere classifiedaccordingto Kenney (1). Au assessmentof strmal fibrousconnectivetissue and focal periglandularfibrosisor fibrotic nests was made. There were no significantdifferencesin luminalepithelialcell heightsor in the occurrenceand severityof stranalfibrousconnectivetissue, focal periglandularfibrosis,or lymphaticlacunaeamong locations(P > 0.051. There was an effect of locationon the occurrenceand severityof inflammation (P < 0.051. If only inflamnation was consideredin categorization, this would have resultedin changingthe categoryin 9 of 60 sanples. There was no increase in tendencyfor inflammtion, fibrosisor lymphaticlacunaeto occur in the horns versus the body of the uterus,nor of the dorsal versus the ventraluterus (P > 0.05). There was no effect (P > 0.05) of type of stain on the abilityto detect incidenceand severityof focal periglandularfibrosis. There was an effect (P < 0.05) of type of stain on the abilityto detect the incidenceand severityof strcmal fibrousconnectivetissue. The use of the trichranestain stied slightlyincreaseddistributionof straml fibrousconnectivetissue.
Keywords: endcmetrim, biopsy,fibrosis,endaaetiitis,lymphaticlacunae
Acknmledgments Supportedin part by BRSG 5-20559awardedby the Bianeilical ResearchSupport Grant Program,Divisionof ResearchResources,NationalInstitutesof Health. 3 current addressis Departmentof VeterinaryMedicineand Surgery,Texas VeterinaryMedicalCenter,Texas A&M University,CollegeStation,M 77843-4475
OCTOBER
1987 VOL. 28 NO. 4
445
THERIOGENOLOGY
Interpretingan endmatrial biopsyhas becane ah integralpart of assessing the potentialfertilityof the brood mare (l-3). Few studieshave reportedon hew representative a single biopsy is of the entire endanetrium. In one study using 16 mares, researcherscanparedsamplestaken fran up to six different locationswithin the uterus (4), and they found that one samplewas generally sufficientfor representingthe mre camkon changes (inflamation,fibrosisand lymphaticlacunae)that cccur in the mdanetrium. Hcwever,many pathologistsand cliniciansalike recumxzndrepeat or nultiplebiopsiesin CategoryIII mares (accordingto Ketmeyl for which there is doubt or cohcernabout the prognosis. While samplesfrom only five mares are includedin this data, our study assesses the accuracywith which a single sampleof uterinetissuecan be used to identify the incidenceand severityof lesionsthroughoutthe entire endanetrim. Widespreadendanetrialfibrcsis,particularlywhen it is periglandular, can depress the abilityof a mre's ehdanstriuin to supporta foal to tenn (l-3). It is cannon practicein sane laboratories to routinelyprepareequine endanetrial specimenswith a connectivetissuestain such as Masson'strichransstain along with the usual hmatoxylin and eosin [H&E)stain to more readilyidentify pathologicendanetrialfibrosis. cu study canparesMasson'strichrarewith hematoxylinand eosin stains for detectingfibrosisin eguine -trial samples.
M?i!rERI?GANDMExioAs Five nonpregnantmares of unknownage and breedinghistorywere used in this study. After either euthanasiaor naturaldeath, their uteri were renovedand ehdanetrialsamplesware taken franeach of12locations: thedorsal aspectof the tip of the left horn (D&B), the dorsal aspect of the left horn (DIE),the dorsal aspect of the right horn (DBH),the dorsal aspectof the tip of the right horn (DTmi),the ventralaspect of the tip of the left horn (VTLJi), the ventral aspect of the left horn (VIE),the ventralaspect of the right horn NBH), the ventralaspect of the tip of the right horn P?lX-I), the ventralaspect of the uterinebody (VB),the dorsal aspect of the uterinebcdy (DB),the ventralaspect of the cervix-uterine body junctiontXB1 and the dorsal aspect of the cervixuterinebcdy junction 0X3). Bmples fran the left and right horns (DLH,DBB, VLH, VBI-I) were taken in the area of the externalbifurcation. Bamplesfran the body of theuterus (VB, DB) were taken approximately one to two inches cranialto the internalcervicalce, while theU3BandJXBsampleswere taken fran the body of ths uterus just anteriorto where it joinedthe cervix. Bach samplewas fixed in Bouin'ssolutionfor 24 h beforetransferto 70% ethanoluntil it was e&added in paraffinand sectionedand stainedwith H&E. A duplicatesectionof each sarrple was also stainedwith Masson'strichranestain. All samplesware interpretedaccordingtoKenmy (1) bycaeprcsector CITLB) slides in a blind manner. In this way, identicalH&E- and trichrane-stained could later be carparedto see if the stain affectedthe interpretation of the amount and severityof endanetrialfibrosis. Fibrosiswas classifiedas either stranalfibrousconnectivetissue or focal periglandularfibrosis. Focal periglandularfibrosiswas expressedas the averagenumber of fibroticgland were ltinal cell heightsin nests per 5.5 n linear field. Also investigated various locations,the distributionand severityof inflamation, freguencyand severityof fibrosisand lymphaticlacunae. Luminalepithelialcell heightwas
446
OCTOBER
1987 VOL. 28 NO. 4
THERIOGENOLOGY
estimated to be cuboidal,low colmnar,nxxkrate columnar,or tallcolumuar accordingto Kenney (1). Additionally,a subjectiveassessmentof glandular densityand depth of gland penetrationinto the laminaprcpriawas made to determineif differencesin these parametersoccurredwith the locationof the sample. Theeffects of locationcm these parametersand the effect of stain cn interpretation of fibrosiswere analyzedby analysisof variancefor split plots in space (5). RENLTS PNDDISCUSSION There were no significantdifferencesin luminalepithelialheight or in the occurrencea~ severityof strcmalfibram connectivetissue,focal periglandular fibrosis,or lymphaticlacunaeamong locations(P > 0.05). Hmaver, the occurrenceand severityof inflamation did vary with location (P < 0.05). If only the inflamation was consideredin arrivingat an overallprognostic CategoryI, II, or III (accordingto Kenney),this weld have resultedin changingthe categoryin 9 out of 60 smples. However,when allpatholcgic changesware taken into considerationin categorizingthe ehdanetrialsamples, the varyingseverityof inflamsation with locationweld only have resultedin changingthe categoryin 1 out of 60 sanples. ,Therewas uo predilection(I,> 0.05) for inflamation, fibrosisor lymphatic lacunaeto occur in the uterinehorns versus the uterinebody, or in the dorsal versus ventraluterus. Lyqhatic lacunaetend to occur more frequentlyin flaccid,enlargeduteri of older mares. Fluid tends to accumulatein the rrore peridant portionsof tissue. Therefore,we were sanswhatsurprisedto see that lymphaticlacuuaedid not appear to occur any nnre frequentlyin ventralsections than in dorsal sections. Hcwever,the eudatrstria of only two of the five mares hadlymphaticlacuuae and than thelacunaeweremild. Perhapstherewould have beet-~ a ventralpatternof preponderance of lymphaticlacunaeif there had been a mare with extensiveendcmatrialcysts and lacunae. We also recognizethat this study used only five mares. Our resultsmight have been differenthad more mares with a broaderspectrumof severityof pathologicchangesbeen included. H-ever, we feel that our findingssupportthose of Bergmanand Kenuey (4) that one biopsywas generallyrepresentative of the entire endanetrium.When there is doubt or concernabout a CategoryIII biopsy,a repeatbiopsy should be taken. Glanddensity was sparse and glanddepthwas shallowin samplestaken near the cervix (Figures1 and 2). This is an importantfindingsince these conditionsare sanetimesinterpretedas pathologicatrophy (61 and are occasionallysuspectedto result fron hormonalimbalances(7). Cliniciansshould be aware that low gland densityor shallowgland depthmay be normal as are pathologicmicroanatcmic cccurrences. In a study on seasonvariationin endmetrial histamorpholcgy, seasonalchangeswere reflectedin end-trial glands and strma that influencedquantitativeassessmentof fibrosis (8). The authorssuggestedthat due to the presenceof normal,physiologicstranaledema observedduring the eetrouscycle, as well as enlargementof nonfibroticglands betweennests, both fibroticand nanfibroticnestingbeccms less frequentduring the physiologicbreedingseason. This informationcould also have teen confoundedby locationof the repeatbiopsies. Biopsiestaken nearer the cervix might have fewer glandularelemants,thereby affectingthe man number of fibrotic glandularnests present. Certainlyany future study of seasonaleffectson the eudcmtrium should take into accountlocationof biopsy.
OCTOBER
1987 VOL. 28 NO. 4
447
THERIOGENOLOGY
Figure 1. Photmicrographof an equine emkmstiial sample taken at the cervix uterinebcdy junction. Notethelajglanddensity and shallcw penetrationof gkmds into the laminapropria.
448
OCTOBER
1987 VOL. 28 NO. 4
THERIOGENOLOGY
sample (samemare as in Figure 2. Photmicrographof an equine *trial Figurel) taken 4 ananteriortothecervix. Glanddensity and penetrationinto the laminapropria is representative of that encounteredin usual biopsies.
OCTOBER
1987 VOL. 28 NO. 4
449
THERIOGENOLOGY
There was no difference(P > 0.05) betweenstains in diagnosingthe incidenceand severityof focal periglahdularfiorosis. However,there was ah effect (P < 0.05) of stain cm the diagnosingof incidenceand severityof stronal fibrousconnectivetissue,with the trichrarts stain resultingin the interpretation of a slight increasein fibrousCOMeCtiVe tissue over that seen with the Ii&E-stained samples. Since there is no data publishedto documentthat ah increasein stranalfibrousconnectivetissue resultsin infertility,the significanceof this data should be interpretedwith caution. Overall,hauever, in cnly two cases would the differencesin anount of strcmalfibrousconnective tissueseen in the trichraua-stained sampleshave resultedin a change in the overallprognostic categoryof the ehdanetrium.We recognizethat the person interpretingthese sampleswas trainedstrictlywith H&E-stainedspecimens. Perhapsthe resultswould have been differentif anotherinterpreterread the samples. Wedo feel it is doubtfulthat trichranestainingof end-trial bicpsiesis of any significantbenefit. REFEREIKES 1.
Kermey,R.M. Cyclic and pathologicchangesof the mare endauetriumas detectedby biopsy,with a note on early enbryonicdeath. J. Am. Vet. Med. Assoc.-:241-262 (1978).
2. Schideler,R.K., Mcchesney,A.E., Voss, J.L. aud Squires,E.L. Relationship of endcmstrialbiopsy and other managementfactorson fertilityof brccdmares. J. Eq. Vet. Sci. 2:5-10 (1982). 3. Neely, D.P. Evaluationand therapyof genitaldisease in the mare. In: Hughes,J.P.,(sd.).Equine Reproduction.veterinaryLearningSystemsCo., II-X=., Izmenceville, Nau Jersey,1983, pp. 39-56. of a uterinebiopsy in the 4. Sergmsn,R.V. and KeMey, R.M. Representativeness mare. Prcc. Am. Assoc. Equine Prcthr.,pp. 355-362 (1975). 5. Steele,R.G.D. and Torrie,J.H. Principlesand Proceduresof Statistics. 1YcGraw-Hill Hz& Co., Inc., New yak, 1960, pp. 247-249. 6. Shideler,R.K., McChesney,A.E. and Voss, J.L. Endanetrialchanges associatedwith infertilityin the mare. Proc. Ann. Mtg. Sac. Therio.,pp. 218-242 (1981). 7. Shideler,R.K. Personalcamnmication,Equine ReprdluctionShort Ccxrse, Fort Collins,Co, January,1983. 8. Gross, T.L. and LeHlauc,M.M. Seasonalvariationof histanorpholcgic featuresof equine endcmstrium. J. Am. Vet. Med. Assoc.-184:1379-1382 (1984).
450
OCTOBER
1987 VOL. 28 NO. 4
INVESTIGATION OF'IHE REPRFSENTATIVENESS OFASlNGLE EN~~ALSAMPLEANDTHEUSEOFTRICHldCMESTAININGTOAID INX-IEDETECI'IONOFENLXMlWRIALFIBROSISINTIiEMARE T.L. Blanchard,1'3 M.C. Garcia:, L.D. Kintner' and R.M. Kenney 1 Hofman Center for ReproductiveStudies New BoltonCenter - Universityof Pennsylvania KennettSquare,PA 91348 and 2veterinaryMedicalDiagnosticLaboratory Universityof Missouri-Colmbia Columbia,MU 65211 Receivedfar publication: JuZy 25, 1986
Accepted: August 5, 1987
The uteri of fivemares were rmovedand er&xnetrialsampleswereprocured fran 12 specificlocationsin the uteri and the saq~leswere processedand duplicatesectionswere stainedwith henatoxylinand eosin (H&E)or Masson's trichrcm stains. The sampleswere interpretedin a blind manner by one person, and pathologicchangeswere classifiedaccordingto Kenney (1). Au assessmentof strmal fibrousconnectivetissue and focal periglandularfibrosisor fibrotic nests was made. There were no significantdifferencesin luminalepithelialcell heightsor in the occurrenceand severityof stranalfibrousconnectivetissue, focal periglandularfibrosis,or lymphaticlacunaeamong locations(P > 0.051. There was an effect of locationon the occurrenceand severityof inflammation (P < 0.051. If only inflamnation was consideredin categorization, this would have resultedin changingthe categoryin 9 of 60 sanples. There was no increase in tendencyfor inflammtion, fibrosisor lymphaticlacunaeto occur in the horns versus the body of the uterus,nor of the dorsal versus the ventraluterus (P > 0.05). There was no effect (P > 0.05) of type of stain on the abilityto detect incidenceand severityof focal periglandularfibrosis. There was an effect (P < 0.05) of type of stain on the abilityto detect the incidenceand severityof strcmal fibrousconnectivetissue. The use of the trichranestain stied slightlyincreaseddistributionof straml fibrousconnectivetissue.
Keywords: endcmetrim, biopsy,fibrosis,endaaetiitis,lymphaticlacunae
Acknmledgments Supportedin part by BRSG 5-20559awardedby the Bianeilical ResearchSupport Grant Program,Divisionof ResearchResources,NationalInstitutesof Health. 3 current addressis Departmentof VeterinaryMedicineand Surgery,Texas VeterinaryMedicalCenter,Texas A&M University,CollegeStation,M 77843-4475
OCTOBER
1987 VOL. 28 NO. 4
445
THERIOGENOLOGY
Interpretingan endmatrial biopsyhas becane ah integralpart of assessing the potentialfertilityof the brood mare (l-3). Few studieshave reportedon hew representative a single biopsy is of the entire endanetrium. In one study using 16 mares, researcherscanparedsamplestaken fran up to six different locationswithin the uterus (4), and they found that one samplewas generally sufficientfor representingthe mre camkon changes (inflamation,fibrosisand lymphaticlacunae)that cccur in the mdanetrium. Hcwever,many pathologistsand cliniciansalike recumxzndrepeat or nultiplebiopsiesin CategoryIII mares (accordingto Ketmeyl for which there is doubt or cohcernabout the prognosis. While samplesfrom only five mares are includedin this data, our study assesses the accuracywith which a single sampleof uterinetissuecan be used to identify the incidenceand severityof lesionsthroughoutthe entire endanetrim. Widespreadendanetrialfibrcsis,particularlywhen it is periglandular, can depress the abilityof a mre's ehdanstriuin to supporta foal to tenn (l-3). It is cannon practicein sane laboratories to routinelyprepareequine endanetrial specimenswith a connectivetissuestain such as Masson'strichransstain along with the usual hmatoxylin and eosin [H&E)stain to more readilyidentify pathologicendanetrialfibrosis. cu study canparesMasson'strichrarewith hematoxylinand eosin stains for detectingfibrosisin eguine -trial samples.
M?i!rERI?GANDMExioAs Five nonpregnantmares of unknownage and breedinghistorywere used in this study. After either euthanasiaor naturaldeath, their uteri were renovedand ehdanetrialsamplesware taken franeach of12locations: thedorsal aspectof the tip of the left horn (D&B), the dorsal aspect of the left horn (DIE),the dorsal aspect of the right horn (DBH),the dorsal aspectof the tip of the right horn (DTmi),the ventralaspect of the tip of the left horn (VTLJi), the ventral aspect of the left horn (VIE),the ventralaspect of the right horn NBH), the ventralaspect of the tip of the right horn P?lX-I), the ventralaspect of the uterinebody (VB),the dorsal aspect of the uterinebcdy (DB),the ventralaspect of the cervix-uterine body junctiontXB1 and the dorsal aspect of the cervixuterinebcdy junction 0X3). Bmples fran the left and right horns (DLH,DBB, VLH, VBI-I) were taken in the area of the externalbifurcation. Bamplesfran the body of theuterus (VB, DB) were taken approximately one to two inches cranialto the internalcervicalce, while theU3BandJXBsampleswere taken fran the body of ths uterus just anteriorto where it joinedthe cervix. Bach samplewas fixed in Bouin'ssolutionfor 24 h beforetransferto 70% ethanoluntil it was e&added in paraffinand sectionedand stainedwith H&E. A duplicatesectionof each sarrple was also stainedwith Masson'strichranestain. All samplesware interpretedaccordingtoKenmy (1) bycaeprcsector CITLB) slides in a blind manner. In this way, identicalH&E- and trichrane-stained could later be carparedto see if the stain affectedthe interpretation of the amount and severityof endanetrialfibrosis. Fibrosiswas classifiedas either stranalfibrousconnectivetissue or focal periglandularfibrosis. Focal periglandularfibrosiswas expressedas the averagenumber of fibroticgland were ltinal cell heightsin nests per 5.5 n linear field. Also investigated various locations,the distributionand severityof inflamation, freguencyand severityof fibrosisand lymphaticlacunae. Luminalepithelialcell heightwas
446
OCTOBER
1987 VOL. 28 NO. 4
THERIOGENOLOGY
estimated to be cuboidal,low colmnar,nxxkrate columnar,or tallcolumuar accordingto Kenney (1). Additionally,a subjectiveassessmentof glandular densityand depth of gland penetrationinto the laminaprcpriawas made to determineif differencesin these parametersoccurredwith the locationof the sample. Theeffects of locationcm these parametersand the effect of stain cn interpretation of fibrosiswere analyzedby analysisof variancefor split plots in space (5). RENLTS PNDDISCUSSION There were no significantdifferencesin luminalepithelialheight or in the occurrencea~ severityof strcmalfibram connectivetissue,focal periglandular fibrosis,or lymphaticlacunaeamong locations(P > 0.05). Hmaver, the occurrenceand severityof inflamation did vary with location (P < 0.05). If only the inflamation was consideredin arrivingat an overallprognostic CategoryI, II, or III (accordingto Kenney),this weld have resultedin changingthe categoryin 9 out of 60 smples. However,when allpatholcgic changesware taken into considerationin categorizingthe ehdanetrialsamples, the varyingseverityof inflamsation with locationweld only have resultedin changingthe categoryin 1 out of 60 sanples. ,Therewas uo predilection(I,> 0.05) for inflamation, fibrosisor lymphatic lacunaeto occur in the uterinehorns versus the uterinebody, or in the dorsal versus ventraluterus. Lyqhatic lacunaetend to occur more frequentlyin flaccid,enlargeduteri of older mares. Fluid tends to accumulatein the rrore peridant portionsof tissue. Therefore,we were sanswhatsurprisedto see that lymphaticlacuuaedid not appear to occur any nnre frequentlyin ventralsections than in dorsal sections. Hcwever,the eudatrstria of only two of the five mares hadlymphaticlacuuae and than thelacunaeweremild. Perhapstherewould have beet-~ a ventralpatternof preponderance of lymphaticlacunaeif there had been a mare with extensiveendcmatrialcysts and lacunae. We also recognizethat this study used only five mares. Our resultsmight have been differenthad more mares with a broaderspectrumof severityof pathologicchangesbeen included. H-ever, we feel that our findingssupportthose of Bergmanand Kenuey (4) that one biopsywas generallyrepresentative of the entire endanetrium.When there is doubt or concernabout a CategoryIII biopsy,a repeatbiopsy should be taken. Glanddensity was sparse and glanddepthwas shallowin samplestaken near the cervix (Figures1 and 2). This is an importantfindingsince these conditionsare sanetimesinterpretedas pathologicatrophy (61 and are occasionallysuspectedto result fron hormonalimbalances(7). Cliniciansshould be aware that low gland densityor shallowgland depthmay be normal as are pathologicmicroanatcmic cccurrences. In a study on seasonvariationin endmetrial histamorpholcgy, seasonalchangeswere reflectedin end-trial glands and strma that influencedquantitativeassessmentof fibrosis (8). The authorssuggestedthat due to the presenceof normal,physiologicstranaledema observedduring the eetrouscycle, as well as enlargementof nonfibroticglands betweennests, both fibroticand nanfibroticnestingbeccms less frequentduring the physiologicbreedingseason. This informationcould also have teen confoundedby locationof the repeatbiopsies. Biopsiestaken nearer the cervix might have fewer glandularelemants,thereby affectingthe man number of fibrotic glandularnests present. Certainlyany future study of seasonaleffectson the eudcmtrium should take into accountlocationof biopsy.
OCTOBER
1987 VOL. 28 NO. 4
447
THERIOGENOLOGY
Figure 1. Photmicrographof an equine emkmstiial sample taken at the cervix uterinebcdy junction. Notethelajglanddensity and shallcw penetrationof gkmds into the laminapropria.
448
OCTOBER
1987 VOL. 28 NO. 4
THERIOGENOLOGY
sample (samemare as in Figure 2. Photmicrographof an equine *trial Figurel) taken 4 ananteriortothecervix. Glanddensity and penetrationinto the laminapropria is representative of that encounteredin usual biopsies.
OCTOBER
1987 VOL. 28 NO. 4
449
THERIOGENOLOGY
There was no difference(P > 0.05) betweenstains in diagnosingthe incidenceand severityof focal periglahdularfiorosis. However,there was ah effect (P < 0.05) of stain cm the diagnosingof incidenceand severityof stronal fibrousconnectivetissue,with the trichrarts stain resultingin the interpretation of a slight increasein fibrousCOMeCtiVe tissue over that seen with the Ii&E-stained samples. Since there is no data publishedto documentthat ah increasein stranalfibrousconnectivetissue resultsin infertility,the significanceof this data should be interpretedwith caution. Overall,hauever, in cnly two cases would the differencesin anount of strcmalfibrousconnective tissueseen in the trichraua-stained sampleshave resultedin a change in the overallprognostic categoryof the ehdanetrium.We recognizethat the person interpretingthese sampleswas trainedstrictlywith H&E-stainedspecimens. Perhapsthe resultswould have been differentif anotherinterpreterread the samples. Wedo feel it is doubtfulthat trichranestainingof end-trial bicpsiesis of any significantbenefit. REFEREIKES 1.
Kermey,R.M. Cyclic and pathologicchangesof the mare endauetriumas detectedby biopsy,with a note on early enbryonicdeath. J. Am. Vet. Med. Assoc.-:241-262 (1978).
2. Schideler,R.K., Mcchesney,A.E., Voss, J.L. aud Squires,E.L. Relationship of endcmstrialbiopsy and other managementfactorson fertilityof brccdmares. J. Eq. Vet. Sci. 2:5-10 (1982). 3. Neely, D.P. Evaluationand therapyof genitaldisease in the mare. In: Hughes,J.P.,(sd.).Equine Reproduction.veterinaryLearningSystemsCo., II-X=., Izmenceville, Nau Jersey,1983, pp. 39-56. of a uterinebiopsy in the 4. Sergmsn,R.V. and KeMey, R.M. Representativeness mare. Prcc. Am. Assoc. Equine Prcthr.,pp. 355-362 (1975). 5. Steele,R.G.D. and Torrie,J.H. Principlesand Proceduresof Statistics. 1YcGraw-Hill Hz& Co., Inc., New yak, 1960, pp. 247-249. 6. Shideler,R.K., McChesney,A.E. and Voss, J.L. Endanetrialchanges associatedwith infertilityin the mare. Proc. Ann. Mtg. Sac. Therio.,pp. 218-242 (1981). 7. Shideler,R.K. Personalcamnmication,Equine ReprdluctionShort Ccxrse, Fort Collins,Co, January,1983. 8. Gross, T.L. and LeHlauc,M.M. Seasonalvariationof histanorpholcgic featuresof equine endcmstrium. J. Am. Vet. Med. Assoc.-184:1379-1382 (1984).
450
OCTOBER
1987 VOL. 28 NO. 4