11

CD56 Expression in Bone and CD56 Expression in Bone and Osteoblastic Neoplasia:Osteoblastic Neoplasia:

A Novel Osteoblast Marker?A Novel Osteoblast Marker?

CD56 Expression in Bone and CD56 Expression in Bone and Osteoblastic Neoplasia:Osteoblastic Neoplasia:

A Novel Osteoblast Marker?A Novel Osteoblast Marker?

D.E. Hughes, R. Deb, D.C.Mangham, V.P. D.E. Hughes, R. Deb, D.C.Mangham, V.P. Sumathi and R.J. GrimerSumathi and R.J. Grimer

Royal Orthopaedic Hospital, Birmingham, Royal Orthopaedic Hospital, Birmingham, U.K.U.K.

D.E. Hughes, R. Deb, D.C.Mangham, V.P. D.E. Hughes, R. Deb, D.C.Mangham, V.P. Sumathi and R.J. GrimerSumathi and R.J. Grimer

Royal Orthopaedic Hospital, Birmingham, Royal Orthopaedic Hospital, Birmingham, U.K.U.K.

22

CD56/NCAMCD56/NCAMCD56/NCAMCD56/NCAMCell adhesion molecule of the Cell adhesion molecule of the

immunoglobulin superfamilyimmunoglobulin superfamilyInvolved in homotypic cell-cell Involved in homotypic cell-cell

interactionsinteractionsPossible role in neural morphogenesisPossible role in neural morphogenesisRestricted tissue distributionRestricted tissue distribution

Cell adhesion molecule of the Cell adhesion molecule of the immunoglobulin superfamilyimmunoglobulin superfamily

Involved in homotypic cell-cell Involved in homotypic cell-cell interactionsinteractions

Possible role in neural morphogenesisPossible role in neural morphogenesisRestricted tissue distributionRestricted tissue distribution

33

CD56 expression in normal CD56 expression in normal tissuestissues

CD56 expression in normal CD56 expression in normal tissuestissues

Neurons, gangliocytesNeurons, gangliocytesPlasma cellsPlasma cellsNatural killer cellsNatural killer cells

Neurons, gangliocytesNeurons, gangliocytesPlasma cellsPlasma cellsNatural killer cellsNatural killer cells

44

CD56 expression in tumoursCD56 expression in tumoursCD56 expression in tumoursCD56 expression in tumours

MyelomaMyelomaNeuroendocrine carcinomasNeuroendocrine carcinomasNeuroblastomaNeuroblastomaPhaeochromocytomaPhaeochromocytomaSome glial tumoursSome glial tumoursNK-type LGLNK-type LGL

MyelomaMyelomaNeuroendocrine carcinomasNeuroendocrine carcinomasNeuroblastomaNeuroblastomaPhaeochromocytomaPhaeochromocytomaSome glial tumoursSome glial tumoursNK-type LGLNK-type LGL

55

Previous evidence of CD56 Previous evidence of CD56 expression in osteoblast expression in osteoblast

lineagelineage

Previous evidence of CD56 Previous evidence of CD56 expression in osteoblast expression in osteoblast

lineagelineage

Expressed by calvarial cells in vitroExpressed by calvarial cells in vitroDescribed in association with CD56 expression Described in association with CD56 expression

in myeloma in osteoblastsin myeloma in osteoblastsReported in osteosarcomasReported in osteosarcomasNot expressed in Ewing’s sarcomaNot expressed in Ewing’s sarcomaNot expressed in chondrosarcomaNot expressed in chondrosarcoma

Expressed by calvarial cells in vitroExpressed by calvarial cells in vitroDescribed in association with CD56 expression Described in association with CD56 expression

in myeloma in osteoblastsin myeloma in osteoblastsReported in osteosarcomasReported in osteosarcomasNot expressed in Ewing’s sarcomaNot expressed in Ewing’s sarcomaNot expressed in chondrosarcomaNot expressed in chondrosarcoma

66

Aim of studyAim of studyAim of studyAim of study

To assess CD56 expression in normal To assess CD56 expression in normal cells of the osteoblast lineagecells of the osteoblast lineage

To assess CD56 expression in tumour-To assess CD56 expression in tumour-like lesions in which bone is formedlike lesions in which bone is formed

To assess CD56 expression in To assess CD56 expression in osteoblastic neoplasmsosteoblastic neoplasms

To assess CD56 expression in normal To assess CD56 expression in normal cells of the osteoblast lineagecells of the osteoblast lineage

To assess CD56 expression in tumour-To assess CD56 expression in tumour-like lesions in which bone is formedlike lesions in which bone is formed

To assess CD56 expression in To assess CD56 expression in osteoblastic neoplasmsosteoblastic neoplasms

77

MethodsMethodsMethodsMethods

Cases selected from Royal Orthopaedic Cases selected from Royal Orthopaedic Hospital paraffin archive to represent a Hospital paraffin archive to represent a broad range of normal bone, reactive broad range of normal bone, reactive conditions, tumour-like conditions and conditions, tumour-like conditions and osteoblastic tumoursosteoblastic tumours

CD56 expression assessed by a standard CD56 expression assessed by a standard immunohistochemical approachimmunohistochemical approach

Cases selected from Royal Orthopaedic Cases selected from Royal Orthopaedic Hospital paraffin archive to represent a Hospital paraffin archive to represent a broad range of normal bone, reactive broad range of normal bone, reactive conditions, tumour-like conditions and conditions, tumour-like conditions and osteoblastic tumoursosteoblastic tumours

CD56 expression assessed by a standard CD56 expression assessed by a standard immunohistochemical approachimmunohistochemical approach

88

ImmunohistochemistryImmunohistochemistryImmunohistochemistryImmunohistochemistryNovacastra anti- Novacastra anti-

CD56 clone 1B6 at CD56 clone 1B6 at 1:501:50

High temperature High temperature antigen retrieval in antigen retrieval in citrate/EDTAcitrate/EDTA

Detection by Detection by Enivision systemEnivision system

Novacastra anti- Novacastra anti- CD56 clone 1B6 at CD56 clone 1B6 at 1:501:50

High temperature High temperature antigen retrieval in antigen retrieval in citrate/EDTAcitrate/EDTA

Detection by Detection by Enivision systemEnivision system

AppendixAppendix

CD56 stainingCD56 stainingof neural cellsof neural cells

99

Case selection – benign Case selection – benign Case selection – benign Case selection – benign Normal/reactive bone (31 cases)Normal/reactive bone (31 cases)

Quiescent boneQuiescent bone Callus, periosteal reactionCallus, periosteal reaction

Benign bone tumours and tumour-like conditions (24 cases)Benign bone tumours and tumour-like conditions (24 cases) Aneurysmal bone cystAneurysmal bone cyst Fibrous dysplasiaFibrous dysplasia Osteofibrous dysplasiaOsteofibrous dysplasia Non-ossifying fibromaNon-ossifying fibroma Desmoplastic fibromaDesmoplastic fibroma Osteoid osteomaOsteoid osteoma OsteoblastomaOsteoblastoma Giant cell tumourGiant cell tumour

Normal/reactive bone (31 cases)Normal/reactive bone (31 cases) Quiescent boneQuiescent bone Callus, periosteal reactionCallus, periosteal reaction

Benign bone tumours and tumour-like conditions (24 cases)Benign bone tumours and tumour-like conditions (24 cases) Aneurysmal bone cystAneurysmal bone cyst Fibrous dysplasiaFibrous dysplasia Osteofibrous dysplasiaOsteofibrous dysplasia Non-ossifying fibromaNon-ossifying fibroma Desmoplastic fibromaDesmoplastic fibroma Osteoid osteomaOsteoid osteoma OsteoblastomaOsteoblastoma Giant cell tumourGiant cell tumour

1010

Case selection – malignantCase selection – malignantCase selection – malignantCase selection – malignantMalignant osteogenic tumours (23 cases)Malignant osteogenic tumours (23 cases)

High grade intramedullary osteosarcoma High grade intramedullary osteosarcoma Telangiectatic osteosarcomaTelangiectatic osteosarcoma Intermediate grade chondroblastic osteosarcomaIntermediate grade chondroblastic osteosarcoma Parosteal osteosarcomaParosteal osteosarcoma Low grade central osteosarcomaLow grade central osteosarcoma Undifferentiated recurrences of known osteosarcomasUndifferentiated recurrences of known osteosarcomas Non bone-forming intra-osseous sarcomas Non bone-forming intra-osseous sarcomas

Malignant osteogenic tumours (23 cases)Malignant osteogenic tumours (23 cases) High grade intramedullary osteosarcoma High grade intramedullary osteosarcoma Telangiectatic osteosarcomaTelangiectatic osteosarcoma Intermediate grade chondroblastic osteosarcomaIntermediate grade chondroblastic osteosarcoma Parosteal osteosarcomaParosteal osteosarcoma Low grade central osteosarcomaLow grade central osteosarcoma Undifferentiated recurrences of known osteosarcomasUndifferentiated recurrences of known osteosarcomas Non bone-forming intra-osseous sarcomas Non bone-forming intra-osseous sarcomas

1111

Results: normal bone (1)Results: normal bone (1)Results: normal bone (1)Results: normal bone (1)

Strong expression inStrong expression inlining cells and osteoblastslining cells and osteoblasts

1212

Results: normal bone (2)Results: normal bone (2)Results: normal bone (2)Results: normal bone (2)

Weak expression inWeak expression instromal cells adjacent stromal cells adjacent to osteoblast seamsto osteoblast seams

Osteocytes Osteocytes PredominantlyPredominantlynegativenegative

1313

Results: callusResults: callusResults: callusResults: callus

No expressionNo expressionin cartilagein cartilage

No expressionNo expressionin fibroblastsin fibroblasts

Strong expressionStrong expressionin osteoblastsin osteoblasts

1414

Results: periosteal reactionResults: periosteal reactionResults: periosteal reactionResults: periosteal reaction

No expression in outerfibrous layer

Strong expressionin bone-forming areas

1515

Results: aneurysmal bone Results: aneurysmal bone cystcyst

Results: aneurysmal bone Results: aneurysmal bone cystcyst

Strong expression onStrong expression onSurface layer of septalSurface layer of septal

structuresstructures

Background stromal Background stromal Population negativePopulation negative

1616

Results: fibrous dysplasiaResults: fibrous dysplasiaResults: fibrous dysplasiaResults: fibrous dysplasia

Spindle cell component positive – Spindle cell component positive – most strongly adjacent to areas most strongly adjacent to areas

of bone formationof bone formation

Similar results in osteofibrous dysplasiaSimilar results in osteofibrous dysplasia

1717

Results: non-ossifying Results: non-ossifying fibromafibroma

Results: non-ossifying Results: non-ossifying fibromafibroma

Some expression Some expression in spindle cells in spindle cells adjacent to boneadjacent to bone

Similar results in desmoplastic fibromaSimilar results in desmoplastic fibroma

1818

Results: osteoid osteomaResults: osteoid osteomaResults: osteoid osteomaResults: osteoid osteoma

Nidus osteoblastsNidus osteoblasts positivepositive

Similar results in osteoblastomaSimilar results in osteoblastoma

1919

Results: giant cell tumourResults: giant cell tumourResults: giant cell tumourResults: giant cell tumour

Focal weak expressionFocal weak expressionby spindle cell componentby spindle cell component

Osteoclasts negativeOsteoclasts negative

2020

Results: high grade Results: high grade osteosarcomaosteosarcoma

Results: high grade Results: high grade osteosarcomaosteosarcoma

Most tumour cells positiveMost tumour cells positive Strong expression inStrong expression inosteoblastic areasosteoblastic areas

Loss of expression inLoss of expression inchondroblastic areaschondroblastic areas

2121

Results: parosteal Results: parosteal osteosarcomaosteosarcoma

Results: parosteal Results: parosteal osteosarcomaosteosarcoma

Variable expression in spindle cellVariable expression in spindle cellcomponent – stronger adjacent to areas of osteoidcomponent – stronger adjacent to areas of osteoid

Similar results in low grade central osteosarcomaSimilar results in low grade central osteosarcoma

2222

Results: telangiectatic Results: telangiectatic osteosarcomaosteosarcoma

Results: telangiectatic Results: telangiectatic osteosarcomaosteosarcoma

Focal expressionFocal expressionIn undifferentiatedIn undifferentiatedTumour cellsTumour cells

2323

Results: undifferentiated Results: undifferentiated recurrence of high grade recurrence of high grade

osteosarcomaosteosarcoma

Results: undifferentiated Results: undifferentiated recurrence of high grade recurrence of high grade

osteosarcomaosteosarcoma

Areas of expression in the Areas of expression in the absence of osteoid formationabsence of osteoid formation

Similar results in some undifferentiated Similar results in some undifferentiated intra-osseous sarcomasintra-osseous sarcomas

2424

Results - summary by cell Results - summary by cell type: non-neoplastictype: non-neoplastic

Results - summary by cell Results - summary by cell type: non-neoplastictype: non-neoplastic

Normal osteoblasts – always positiveNormal osteoblasts – always positiveNormal osteocytes – mostly negativeNormal osteocytes – mostly negativeNormal periosteal spindle cells – positive Normal periosteal spindle cells – positive

in areas of active bone formationin areas of active bone formationNormal bone marrow stromal cells - Normal bone marrow stromal cells -

positive in areas of active bone formationpositive in areas of active bone formation

Normal osteoblasts – always positiveNormal osteoblasts – always positiveNormal osteocytes – mostly negativeNormal osteocytes – mostly negativeNormal periosteal spindle cells – positive Normal periosteal spindle cells – positive

in areas of active bone formationin areas of active bone formationNormal bone marrow stromal cells - Normal bone marrow stromal cells -

positive in areas of active bone formationpositive in areas of active bone formation

2525

Results - summary by cell type: Results - summary by cell type: neoplasticneoplastic

Results - summary by cell type: Results - summary by cell type: neoplasticneoplastic

Neoplastic osteoblasts – consistently positiveNeoplastic osteoblasts – consistently positive Neoplastic osteocytes – variably positiveNeoplastic osteocytes – variably positive Neoplastic spindle cells with osteogenic Neoplastic spindle cells with osteogenic

potential – variably positivepotential – variably positive Neoplastic spindle cells without evidence of Neoplastic spindle cells without evidence of

osteogenic potential – sometimes positiveosteogenic potential – sometimes positive

Neoplastic osteoblasts – consistently positiveNeoplastic osteoblasts – consistently positive Neoplastic osteocytes – variably positiveNeoplastic osteocytes – variably positive Neoplastic spindle cells with osteogenic Neoplastic spindle cells with osteogenic

potential – variably positivepotential – variably positive Neoplastic spindle cells without evidence of Neoplastic spindle cells without evidence of

osteogenic potential – sometimes positiveosteogenic potential – sometimes positive

2626

ConclusionsConclusionsConclusionsConclusionsCD56 is consistently strongly expressed CD56 is consistently strongly expressed

by normal and neoplastic osteoblastsby normal and neoplastic osteoblastsCD56 expression also occurs in non-CD56 expression also occurs in non-

osteogenic spindle cells in bone and bone osteogenic spindle cells in bone and bone lesions – lesions – it may not be a specific marker it may not be a specific marker of osteoblastic potential for diagnostic of osteoblastic potential for diagnostic purposespurposes

CD56 may be a biologically important CD56 may be a biologically important molecule in osteoblast functionmolecule in osteoblast function

CD56 is consistently strongly expressed CD56 is consistently strongly expressed by normal and neoplastic osteoblastsby normal and neoplastic osteoblasts

CD56 expression also occurs in non-CD56 expression also occurs in non-osteogenic spindle cells in bone and bone osteogenic spindle cells in bone and bone lesions – lesions – it may not be a specific marker it may not be a specific marker of osteoblastic potential for diagnostic of osteoblastic potential for diagnostic purposespurposes

CD56 may be a biologically important CD56 may be a biologically important molecule in osteoblast functionmolecule in osteoblast function