**6.2 Electron microscopic examination**

304 Recent Advances in Arthroplasty

Arbitrary values obtained were compared with a series of standard solutions (Sigma-

Joint fluid malondialdehyde levels were determined by the modified method of Yagi (Yagi, 1982). The samples were read with fluorometric detection at 515/553 nm. As a standard solution Tetraetoxypropane in concentration of 0.1 μmol/L was used. Results were

Hydroxyproline content was evaluated according to the method of Reddy & Enwemeka (Reddy & Enwemeka, 1996). Absorbance of each sample was read at 550 nm using HPLC (Spectrochrom). Serial dilutions of commercial pure hydroxyproline (Sigma-Aldrich) were used as standard. All samples were assayed in duplicates. Results were expressed as mg/g

Elemental concentrations were determined by Inductively Coupled Plasma-Mass Spectrometry (HP-4500; Agilent Technologies, Waldbronn, Germany). Standard reference material was obtained from NIST (RF 1577b, NIST, Gaithersburg, MD). Values were

Histological examination of the periprosthetic tissues showed large amounts of metal and polyethylene debris and a nonspecific chronic inflammatory reaction. Wear debris, macrophages that had phagocytosed small metal and polyethylene particles, fibroid necrosis, and proliferation of capillaries were seen more commonly in granulomas (Fig. 1).

Fig. 1. Polyethylene and metal particles in the interstitium, giant cells and macrophages

Aldrich, St. Louis, MO). Results were expressed as nmol/mg hydroxyproline.

**5.6.2 Malondialdehyde determination in pseudojoint fluid** 

expressed as nmol/L.

wet tissue.

**6. Results** 

**5.7 Collagen determination** 

**5.8 Metal particle analysis** 

measured in μg/g wet weight.

**6.1 Histological examination** 

(Perle's iron stain, x500)

Analysis of selected sections from ten representative cases by electron microscopy established damaged collagen fibers and presence of collagen cross-links. Their relative

Fig. 3. Electron micrograph (x39 000) showing periprosthetic tissue taken from stable cementless hip replacement 86 months after implantation. The picture shows damaged collagen fibers (open arrows) with numerous cross-links (black arrows).

Evidence Linking Elevated Oxidative Stress and Aseptic Loosening of Hip Arthroplasty 307

wear, showed higher oxidative stress in the two groups. The mean MDA value of the 28 patients in Group I was 0.052 nmol/mg (±0.09 SD), and of the 12 patients with high rate of wear and osteolysis in Group II, 0.031 nmol/mg (±0.014 SD) (p=0.74). MDA levels in the control tissues (0.009 nmol/mg, ±0.0093 SD) were significantly lower than those in Group I (p<0.0001) and Group II (p<0.0001). Figure 5 shows the comparisons between the two

Fig. 5. The concentration of lipid peroxidation product, malondialdehyde (MDA), in

did not correlate with pelvic osteolysis or time elapsed from previous surgery.

**6.4.2 Malondialdehyde determination in pseudosynovial fluid** 

periprosthetic tissues and controls. Values were measured in nmol/mg hydroxyproline. The lower and upper lines in the boxes represent the 25th and 75th percentiles, respectively,

Determination of oxidative stress assessed by level of lipid peroxidation product MDA in pseudojoint fluid showed higher oxidative stress in revision cases. The mean MDA value of the 18 patients with loose hip prostheses was 27.5 nmol/L (±17.6 SD, range 13.5 to 82.9). MDA level in the pseudojoint fluid from controls was significantly lower – 14.9 nmol/L (±4.5 SD, range 10.7 to 28.9) (p=0.001). Figure 6 shows the comparison between patients and controls in graphical form. Although not significantly, MDA level correlated moderately with linear polyethylene wear and grade of femoral osteolysis (Spearman's rho=0.321 and rho=0.315, respectively). Oxidative stress measured by MDA level in pseudosynovial fluid

groups and controls in graphical form.

\*p<0.0001 in comparison with controls.

with the median marked in the box.

abundance compared to control samples taken from fascia lata supported the proposal that these findings were a result of ROS damage (Fig. 3). Wear debris inclusions were demonstrated in the various cells (Fig. 4).

Fig. 4. Electron micrograph (x12 000) showing giant cell with ingested polyethylene (open arrow) and metal (black arrow) particles.
