Bone morphogenetic protein-2 (BMP-2) and transforming growth factor-beta1 (TGF-beta1) alter connexin 43 phosphorylation in MC3T3-E1 Cells.

BACKGROUND: Bone morphogenetic proteins (BMPs) and transforming growth factor-betas (TGF-betas) are important regulators of bone repair and regeneration. BMP-2 and TGF-beta1 have been shown to inhibit gap junctional intercellular communication (GJIC) in MC3T3-E1 cells. Connexin 43 (Cx43) has been shown to mediate GJIC in osteoblasts and it is the predominant gap junctional protein expressed in these murine osteoblast-like cells. We examined the expression, phosphorylation, and subcellular localization of Cx43 after treatment with BMP-2 or TGF-beta1 to investigate a possible mechanism for the inhibition of GJIC. RESULTS: Northern blot analysis revealed no detectable change in the expression of Cx43 mRNA. Western blot analysis demonstrated no significant change in the expression of total Cx43 protein. However, significantly higher ratios of unphosphorylated vs. phosphorylated forms of Cx43 were detected after BMP-2 or TGF-beta1 treatment. Immunofluorescence and cell protein fractionation revealed no detectable change in the localization of Cx43 between the cytosol and plasma membrane. CONCLUSIONS: BMP-2 and TGF-beta1 do not alter expression of Cx43 at the mRNA or protein level. BMP-2 and TGF-beta1 may inhibit GJIC by decreasing the phosphorylated form of Cx43 in MC3T3-E1 cells.

Well-differentiated liposarcoma (atypical lipoma) of the lower extremity in a patient with bilateral leg lipodystrophy and lymphedema.

We present a report of a patient with bilateral lower extremity lipodystrophy and lymphedema who underwent excision of a large extremity liposarcoma. Total excision of the tumor was performed with no evidence of recurrence to date. The natural history, characteristics, and management of this tumor are discussed. A high index of suspicion and awareness among surgeons and pathologists should allow accurate diagnosis and treatment of this condition.

Staged skin and subcutaneous excision for lymphedema: a favorable report of long-term results.

Numerous surgical procedures have been proposed for the management of lymphedema. The postoperative results vary, and unfortunately none of the procedures are curative. As a result, some degree of recurrence of leg edema is seen in all patients postoperatively. Reported here is a long-term follow-up of patients with lower extremity lymphedema managed by skin and subcutaneous tissue excision. Thirty-eight patients (6 male; 32 female) with lower extremity lymphedema have been followed up for an average of 14 (3 to 27) years after staged subcutaneous excisions performed beneath skin flaps. Seven patients had been treated previously by other procedures. Of the 38 lymphedema patients, 10 patients developed edema after pelvic or groin ablative surgery, radiation therapy, or both. Results were documented by various methods: physical examination, circumferential measurements, volume displacement, serial photography, lymphoscintigraphy, and patient survey. Of these, it is believed that photographs are the easiest and as representative as any other method, all of which have great variability. Of the 38 patients, 30 patients had significant and long-lasting reduction in extremity size associated with improved function and extremity contour. Episodes of recurrent cellulitis were reduced or completely eliminated. No differences in the long-term results were seen in patients with acquired as opposed to congenital lymphedema. Men did not have as much improvement as women. Two patients had no change in leg swelling, and six patients (three men) had progressive swelling after surgery. Partial wound separation occurred immediately postoperatively in one patient, and three patients had loss (less than 2 cm) of the skin flap, all in the ankle region. None of these instances required further surgery, and no other significant complications were encountered. Staged skin and subcutaneous excision beneath skin flaps appears to provide long-lasting improvement for lower extremity lymphedema, regardless of cause, in the majority of patients treated.

The influence of time on human breast capsule histology: smooth and textured silicone-surfaced implants.

Although the histology of capsular tissue is well described in the literature, most studies in humans do not correlate histologic findings with implant age (number of years an implant was in place before sampling). As such, questions regarding the long-term histology in humans remain. The microanatomy of 93 human periprosthetic capsular tissues surrounding 22 textured and 71 smooth silicone-surfaced prostheses was studied. The implants were divided into two groups according to the time between implantation and capsulectomy: between 0 and 5 years or more than 5 years. Hematoxylin and eosin and Masson trichrome-stained sections were analyzed by light microscopy, with and without polarization. Eighteen of the textured implants contained silicone gel and four contained saline. Sixty of the smooth implants contained silicone gel, eight contained saline, and in three, the filler type was not known. For the majority of patients, surgery was performed for augmentation mammaplasty, and the implants were removed because of capsular contracture. The following histologic features were assessed: synovial-like metaplasia, villous hyperplasia, density of the collagenous capsule, alignment of collagen fibers within the capsule, and the presence of foreign material and of a foreign body reaction. The following trends were observed. In smooth implants, increasing implant duration was associated with a decrease in the presence of synovial-like metaplasia (p = 0.003) and villous hyperplasia; there was no significant difference in the presence of a dense collagenous capsule, the orientation of collagen fibers, or the presence of a foreign body reaction. An increase was observed in the presence of foreign material (p = 0.01). In textured implants, increasing implant duration was associated with a decrease in the presence of synovial-like metaplasia, villous hyperplasia (p = 0.003), dense collagenous architecture, and parallel orientation of collagen fibers (p = 0.017). An increase in the presence of a foreign body type reaction and foreign material (p = 0.024) was observed. In comparing textured and smooth-surfaced implants, synovial-like metaplasia was observed more often in the textured group, both at 0 to 5 years (p = 0.01) and at greater than 5 years (p < 0.01). Textured implants more often had villous hyperplasia at 0 to 5 years (p = 0.03) but not beyond 5 years. Smooth implants more often had a dense collagenous capsule than textured implants after 5 years. No significant difference was seen in the orientation of collagen fibers in capsules around smooth and textured implants at 0 to 5 years. After 5 years, the incidence of capsules with collagen fibers arranged parallel to the implant surface was significantly greater in the smooth group than in the textured group (p = 0.01). The presence of a foreign body type reaction was seen more often in the textured group between 0 and 5 years (p = 0.01) and at greater than 5 years (p < 0.01), and the presence of foreign material was more often seen in the textured group between 0 and 5 years (p = 0.06) and at greater than 5 years (p < 0.01). In summary, the cytologic changes around implants seem to be dynamic in nature, and implantation duration and shell type play a significant role. Synovial-like metaplasia, villous hyperplasia, and foreign material were more often observed in the textured group within the 0 to 5 year interval. Beyond 5 years, synovial-like metaplasia, a foreign body type reaction, and foreign material were more often observed in the textured group. Differences in the density of collagenous capsules were not significant at any time point, and collagen fibers oriented parallel to the implant surface were more often observed in the smooth group after 5 years. The significance of these findings awaits further investigation.

Persistent neutrophil (PMN) activation 24 hr after ischemia and reperfusion.

Neutrophils have been implicated as mediators of the reperfusion injury which occurs in skeletal muscle after ischemia. The purpose of this study is to determine if PMN chemotaxis occur 24 hr after an ischemic injury followed by reperfusion. Rabbit hindlimbs were made ischemic by clamping the right iliac and femoral arteries for 2 or 3 hr after ligating all pelvic collaterals. The limb was then reperfused by removing the clamps. After 24 hr of reperfusion, blood samples were drawn from the carotid artery and right iliac vein. Serum and PMN were isolated from each blood sample. Chemotaxis, as determined by migration across a filter, was measured in each group. One-way analysis of variance demonstrated a significant increase in chemotaxis in PMN isolated from the venous effluent of hind limbs which had undergone 3 hr of ischemia and 24 hr of reperfusion. PMN isolated from the carotid artery of the same animals had significantly less chemotactic activity. There were no significant differences from control in PMN chemotaxis in specimens from sham animals or animals exposed to 2 hr of ischemia and 24 hr of reperfusion. Those rabbits which had undergone 3 hr of ischemia had a clinically stiff and edematous hindlimb, whereas those which had undergone 2 hr of ischemia and the sham animals had a normal hindlimb after 24 hr of reperfusion. PMN isolated from the venous effluent of an ischemic limb after 3 hr of ischemia and 24 hr of reperfusion demonstrated increased chemotaxis while systemic arterial PMN from the same animals showed no more chemotaxis than control cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Edwardsiella tarda in freshwater catfish and their environment.

Edwardsiella tarda was isolated from 47, 88, and 79% of skin, visceral, and dressed-fish samples, respectively. This species was also isolated from 30% of imported dressed fish, 75% of catfish pond water samples, 64% of catfish pond mud samples, and 100% of frogs, turtles, and crayfish from catfish ponds. The incidence of Edwardsiella increased during the summer months, as water temperatures increased. Of several isolation media evaluated, the most effective was selective enrichment in double-strength Salmonella-Shigella broth and subsequent plating on single-strength Samonella-Shigella agar. The significance of the incidence of Edwardsiella in catfish, catfish disease, and public health could not be substantiated.

Serial passage of MC3T3-E1 cells alters osteoblastic function and responsiveness to transforming growth factor-beta1 and bone morphogenetic protein-2.

The murine-derived clonal MC3T3-E1 cell is a well-studied osteoblast-like cell line. To understand the effects of serial passages on its cellular function, we examined changes in cell morphology, gap junctional intercellular communication (GJIC), proliferation, and osteoblastic function between early passage (<20) and late passage (>65) cells. MC3T3-E1 cells developed an elongated, spindle shape after multiple passages. Intercellular communication decreased significantly (33%) in late vs. early passage cells. Transforming growth factor-beta1 (TGF-beta1) stimulated cell proliferation in early passage cells and induced c-fos expression, while it inhibited proliferation in late passage cells. Using alkaline phosphatase (ALP) activity and osteocalcin (OC) secretion as markers for osteoblastic function and differentiation, we demonstrated that both markers were significantly reduced after multiple cell passages. Bone morphogenetic protein-2 (BMP-2) significantly enhanced ALP activity and OC secretion in early passage cells while TGF-beta1 exerted an opposite effect. Both BMP-2 and TGF-beta1 had minimal effects on late passage cells. We conclude that serial passage alters MC3T3-E1 cell morphology, and significantly diminishes GJIC, osteoblastic function, TGF-beta1-mediated cell proliferation, and responsiveness to TGF-beta1 and BMP-2. Cell passage numbers should be clearly defined in functional studies involving MC3T3-E1 cells.