Results: 4

1.
Fig. 4.

Fig. 4. From: RHAMM, a receptor for hyaluronan-mediated motility, compensates for CD44 in inflamed CD44-knockout mice: A different interpretation of redundancy.

CIA in CD44-deficient mice is sensitive to treatment with anti-RHAMM mAb or soluble rRHAMM. CD44+/+ (AC) and CD44–/– (DF) DBA/1 mice, treated twice with type II collagen to induce CIA, were administered 150 μg of anti-RHAMM mAb or PBS (A and D, eight mice in each group), 150 μg of anti-RHAMM mAb or isotype-matched IgG1 (B and E, five mice in each group), using the protocol described in Fig. 3, or 1 mg/kg soluble mouse GST-rRHAMM or GST (C and F, eight mice in each group), administered the day of the first collagen injection and then twice a week for 5 weeks. Arthritis development was monitored for 8–10 days by measuring paw swelling. *, P < 0.05; **, P < 0.01, by Student's t test for unpaired values. The results are expressed as the mean ± SE.

Shlomo Nedvetzki, et al. Proc Natl Acad Sci U S A. 2004 December 28;101(52):18081-18086.
2.
Fig. 2.

Fig. 2. From: RHAMM, a receptor for hyaluronan-mediated motility, compensates for CD44 in inflamed CD44-knockout mice: A different interpretation of redundancy.

The compensating molecule in CD44-deficient mice recognizes HA. (A and B) CIA in CD44-deficient mice is refractory to treatment with anti-CD44 mAb. CD44–/– (A) and CD44+/+ and CD44+/– (B) DBA/1 mice (five in each group), showing symptoms of arthritis after two injections of type II collagen, were treated upon disease onset and then every other day for 10 days with 150 μg of IM7.8.1 anti-CD44 mAb or isotype-matched control mAb (see symbols). Arthritis development was monitored for 10 days by measuring paw swelling. (C and D) CIA in CD44-deficient mice is sensitive to enzymatic treatment with hyaluronidase. WT (C and D Left), heterozygous (D Center), and CD44-deficient (D Right) DBA/1 mice (five in each group), in which CIA was induced after two doses of type II collagen, were treated at the initiation phase of the disease (day 0) and then every other day for 5 weeks with 20 units of hyaluronidase (♦), 10 units (equivalent specific activity) of heparinase (▴), or PBS (▪). Arthritis development was monitored from disease onset and then for 12–14 days by measuring paw swelling. *, P < 0.05; **, P < 0.01, by Student's t test for unpaired values. The results are expressed as the mean ± SE.

Shlomo Nedvetzki, et al. Proc Natl Acad Sci U S A. 2004 December 28;101(52):18081-18086.
3.
Fig. 3.

Fig. 3. From: RHAMM, a receptor for hyaluronan-mediated motility, compensates for CD44 in inflamed CD44-knockout mice: A different interpretation of redundancy.

Expression and migratory function of RHAMM in spleen leukocytes of arthritic and nonarthritic CD44-deficient and WT mice. (A) Western blot analysis. Extracts of spleen leukocytes from arthritic (CIA) and nonarthritic CD44+/+, CD44+/–, and CD44–/– DBA/1 mice, treated twice with collagen, were analyzed by Western blot using polyclonal anti-RHAMM antibody. (B) Flow cytometry analysis. Cell surface RHAMM was assessed on spleen leukocytes (Left) from arthritic (CIA) or nonarthritic CD44+/+ and CD44–/– DBA/1 mice or on joint-infiltrating cells derived from arthritic CD44+/+ and CD44–/– mice (Right) by measuring the binding of polyclonal anti-RHAMM antibody to the cells. The binding of anti-RHAMM antibody was identified with anti-rabbit Ig conjugated to fluorescein. The black line in each histogram shows the nonspecific binding of second antibody alone. (C) Enhanced transwell migration of CD44-deficient spleen leukocytes. Spleen leukocytes derived from arthritic (CIA +/+ and CIA –/–) and nonarthritic (+/+ and –/–), WT (+/+), and CD44-deficient (–/–) DBA/1 mice were subjected to the transwell migration assay. The spleen leukocytes were analyzed for their ability to cross filters, noncoated (filled bars) or coated (empty + filled bars) with HA, toward conditioned fibroblast medium that served as chemoattractant. The number of cells that traversed the filter are shown on the y axis. (D and E) Inhibition of transwell migration of spleen leukocytes from arthritic CD44-deficient mice by anti-RHAMM antibody. Transwell migration (across HA-coated filters) of arthritic (–/– CIA; +/+ CIA) and nonarthritic (–/–; +/+) spleen leukocytes from CD44-deficient (D) and WT (E) mice was analyzed as described in C. The assay was performed in the presence of medium (UT, untreated), anti-RHAMM polyclonal antibody (Anti-RHAMM), preimmune rabbit serum (serum), and IM7.8.1 anti-CD44 mAb and isotype-matched control mAb (4D2). Statistical analysis was done by Student's t test for unpaired values. The results are expressed as the mean ± SE.

Shlomo Nedvetzki, et al. Proc Natl Acad Sci U S A. 2004 December 28;101(52):18081-18086.
4.
Fig. 1.

Fig. 1. From: RHAMM, a receptor for hyaluronan-mediated motility, compensates for CD44 in inflamed CD44-knockout mice: A different interpretation of redundancy.

Analysis of CIA in CD44-deficient mice. (A) Infiltrating leukocytes rather than local factors determine the nature of cell surface CD44. Suspensions of spleen leukocytes from CD44–/– mice were infused i.v. into irradiated CD44+/+ mice, and those of CD44+/+ mice were infused i.v. into irradiated CD44–/– mice. All of the recipient mice (five in each group) were immunized with a single dose of type II collagen. CD44 expression on spleen cells of a representative donor (+/+; –/–) or recipient (+/+ → –/–; –/– → +/+) was analyzed by flow cytometry, and the time of arthritis onset (day ± SD) was recorded for each mouse (see text). The binding of IM7.8.1 anti-pan CD44 mAb to spleen leukocytes was detected by fluorescence-labeled anti-mouse Ig antibody. ImC, Ig (secondary antibody) control. (B and C) Histopathological analysis: CIA in CD44-deficient mice is more aggravated than in WT mice. CIA was generated after injection of two doses of type II collagen. Tarsal/metatarsal joints were removed 14 days after CIA onset, decalcified, fixed, and stained with hematoxylin and eosin. (B) WT mice show severe erosion of bone (BO) and cartilage (CA). Some cartilage remains intact, the joint space (JS) is still evident, and the joint architecture is partially maintained. (C) CD44-deficient mice show severe erosion of bone and complete erosion of cartilage. The entire joint architecture (including joint space) has been obliterated. (Inset) Infiltration of a large number of polymorphonuclear leukocytes into the joint space of CD44-deficient mice. The joint was removed at disease onset, and the section was stained with safranin-o after fixation and decalcification. BM, bone marrow. (D) Histochemical photoscanning analysis: Enhanced accumulation of HA in joint tissues of arthritic CD44–/– mice. Tarsal/metatarsal joints were removed 14 days after the onset of CIA in arthritic WT (+/+ CIA) and arthritic CD44-deficient (–/– CIA) mice, subjected (+Hdase) or not subjected (–Hdase) to hyaluronidase treatment, as described. HA accumulation in tissue was identified by a biotinylated HABP and avidin-biotin peroxidase detection system. The intensity of HA accumulation in the tissue was analyzed by photoscanning, as described in Materials and Methods.

Shlomo Nedvetzki, et al. Proc Natl Acad Sci U S A. 2004 December 28;101(52):18081-18086.

Supplemental Content

Recent activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...
Write to the Help Desk