Biogerontology

, Volume 8, Issue 5, pp 483–498 | Cite as

Methodological models for in vitro amplification and maintenance of human articular chondrocytes from elderly patients

  • Anna Maria Carossino
  • Raffaella Recenti
  • Roberto Carossino
  • Elisabetta Piscitelli
  • Alessia Gozzini
  • Valentina Martineti
  • Carmelo Mavilia
  • Alessandro Franchi
  • Daniele Danielli
  • Paolo Aglietti
  • Antonio Ciardullo
  • Gianna Galli
  • Isabella Tognarini
  • Alberto Moggi Pignone
  • Mario Cagnoni
  • Maria Luisa Brandi
Research Article

Abstract

Articular cartilage defects, an exceedingly common problem closely correlated with advancing age, is characterized by lack of spontaneous resolution because of the limited regenerative capacity of adult articular chondrocytes. Medical and surgical therapies yield unsatisfactory short-lasting results. Recently, cultured autologous chondrocytes have been proposed as a source to promote repair of deep cartilage defects. Despite encouraging preliminary results, this approach is not yet routinely applicable in clinical practice, but for young patients. One critical points is the isolation and ex vivo expansion of large enough number of differentiated articular chondrocytes. In general, human articular chondrocytes grown in monolayer cultures tend to undergo dedifferentiation. This reversible process produces morphological changes by which cells acquire fibroblast-like features, loosing typical functional characteristics, such as the ability to synthesize type II collagen. The aim of this study was to isolate human articular chondrocytes from elderly patients and to carefully characterize their morphological, proliferative, and differentiative features. Cells were morphologically analyzed by optic and transmission electron microscopy (TEM). Production of periodic acid-schiff (PAS)-positive cellular products and of type II collagen mRNA was monitored at different cellular passages. Typical chondrocytic characteristics were also studied in a suspension culture system with cells encapsulated in alginate-polylysine-alginate (APA) membranes. Results showed that human articular chondrocytes can be expanded in monolayers for several passages, and then microencapsulated, retaining their morphological and functional characteristics. The results obtained could contribute to optimize expansion and redifferentiation sequences for applying cartilage tissue engineering in the elderly patients.

Keywords

Human articular chondrocytes Type I collagen Type II collagen Alginate Cell therapy 

Abbreviations

ACTB

Beta actin

AGC1

Aggrecan 1

AMV

Avian Myeloblastosis Virus

APA

Alginate-polylysine-alginate

BGN

Biglycan

CHES

2-N-cyclohexylaminoethane sulfonic acid

COLIA1

Type I alpha 1 collagen

COLIIA1

Type II alpha 1 collagen

COLXA1

Type X alpha 1 collagen

DCN

Decorin

D-MEM

Dulbecco’s modified Eagle’s medium

DT

Doubling time

FCS

Fetal calf serum

GAGs

Glycosaminoglycans

GAPDH

Glyceraldehyde-3-phosphate dehydrogenase

OA

Osteoarthritis

PAS

Periodic acid-schiff

PBS

Phosphate buffer saline

PCR

Polymerase-chain reaction

RT

Real time

SDS-PAGE

sodium dodecyl sulfate polyacrylamide gel electrophoresis

TEM

Transmission electron microscopy

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Copyright information

© Springer Science+Business Media, Inc. 2007

Authors and Affiliations

  • Anna Maria Carossino
    • 1
  • Raffaella Recenti
    • 1
  • Roberto Carossino
    • 1
  • Elisabetta Piscitelli
    • 1
  • Alessia Gozzini
    • 1
  • Valentina Martineti
    • 1
    • 4
  • Carmelo Mavilia
    • 4
  • Alessandro Franchi
    • 2
  • Daniele Danielli
    • 2
  • Paolo Aglietti
    • 3
  • Antonio Ciardullo
    • 3
  • Gianna Galli
    • 1
  • Isabella Tognarini
    • 1
  • Alberto Moggi Pignone
    • 1
  • Mario Cagnoni
    • 1
  • Maria Luisa Brandi
    • 1
    • 4
  1. 1.Departments of Internal MedicineUniversity of FlorenceFlorenceItaly
  2. 2.Departments of Human Pathology and OncologyUniversity of FlorenceFlorenceItaly
  3. 3.Departments of OrthopedicsUniversity of FlorenceFlorenceItaly
  4. 4.DeGene Spin-offUniversity of FlorenceFlorenceItaly

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