Cryopreservation of human adipose tissues.

Scientific studies on cryopreservation of adipose tissues have seldom been performed. The purpose of our present study is conducted both in vitro and in vivo to develop a novel cryopreservation method that can be used successfully for long-term preservation of human adipose tissues for possible future clinical application. In this study, samples of adipose aspirates were obtained from 36 adult white female patients after liposuction and collected from the middle layer after centrifugation. In the in vitro study, suitable cryoprotectant agents (CPAs) and their concentrations and possible combinations were selected from our preliminary experiment. A combination of dimethyl sulfoxide (Me(2)SO) and trehalose as CPA with the optimal concentration (0.5M Me(2)SO and 0.2M trehalose) was chosen and then used throughout the study. In addition, maximal recovery of adipose tissues was achieved after cryopreservation using slow cooling without seeding (1-2 degrees C/min to -30 degrees C, followed by plunging to -196 degrees C for storage) and fast warming (in 40 degrees C water bath, averaging 35 degrees C/min). Fresh adipose aspirates (Group 1), cryopreserved adipose aspirates without CPAs (Group 2), or cryopreserved adipose aspirates with CPAs (Group 3) were evaluated by integrated adipocyte counts and histology. In the in vivo study, fresh adipose aspirates (Group 1), cryopreserved adipose aspirates without CPAs (Group 2), or cryopreserved adipose aspirates with CPAs (Group 3) were injected into a nude mouse. The retained adipose aspirates (fat grafts) were harvested in each animal at 4 months and their weight, volume, and histology was assessed. In the in vitro study, significantly higher integrated viable adipocyte count (2.06+/-0.54 x 10(6)mL(-1) vs. 1.07+/-0.41 x 10(6)mL(-1), p<0.0011) of adipose aspirates was found in Group 3 compared with Group 2. Group 3 had only a marginally lower integrated viable adipocyte count compared with Group 1 (2.06+/-0.54 x 10(6)mL(-1) vs. 2.57+/-0.56 x 10(6)mL(-1), p=0.083). Histologically, more tissue shrinkage was evident in Group 2 compared with Group 3. In the in vivo study, various degrees of absorption of injected fat grafts were seen in all 3 groups. However, Group 3 had significantly more retained weight and volume of the injected fat grafts than Group 2 (both p<0.0001) but had significantly less retained weight and volume than Group 3 (weight, p=0.009178; volume, p=0.007836). Histologically, a large amount of tissue fibrosis was seen in Group 2, and reasonably well maintained fatty tissue with only a small amount of tissue fibrosis was seen in Group 3. The results from the present in vitro and in vivo studies, for the first time, demonstrate that our preferred cryopreservation method, the combination of 0.5M Me(2)SO and 0.2M trehalose as CPA in addition to the controlled slow cooling and fast rewarming protocol, appears to provide the maximum recovered results in cryopreservation of human adipose tissues and may become a real option after further refinements for cryopreservation of human adipose aspirates in a clinical setting.

Enhancement of bone growth by sustained delivery of recombinant human bone morphogenetic protein-2 in a polymeric matrix.

PURPOSE: The purpose of this study was to develop a polymeric sustained delivery system for recombinant human bone morphogenetic protein-2 (BMP-2) and to evaluate local bone growth induced by the sustained release of BMP-2 in an animal model. METHODS: BMP-2 was incorporated in biodegradable poly(D,L-lactide-co-glycolide) (PLGA) microspheres to obtain different release rates. Two sustained and an immediate release implants were produced by suspending the BMP-2 loaded PLGA microspheres in aqueous sodium carboxymethylcellulose (CMC), lyophilizing, and cutting the dried materials to the size of the animal bone defects. The local in vivo release at the implantation site in rat calvarial defects was determined by gamma scintigraphy using radiolabeled BMP-2. The local bone induction in the critical size of rabbit calvarial defects was evaluated six weeks post implantation. RESULTS: The immediate release implant showed about 65% initial drug release within 24 h and the remaining BMP-2 quickly exhausted from the implantation site within 7 days. The sustained release implants, showing 45-55% initial release followed by a prolonged release for 21 days, released a greater amount of BMP-2 at the implantation site and maintained higher serum BMP-2 for the longer period of time compared to the immediate release implant. Significant bone growth was observed in all BMP-2 treated defects while the defects without treatment or with BMP-2-free implant showed minimal bone healing. 75-79% of rabbit calvarial defect area was healed with newly induced bone matrix by the sustained release implants in 6 weeks as compared to 45% recovery from the immediate release implant. CONCLUSION: The sustained delivery of BMP-2 based on the biodegradable PLGA microsphere system resulted in faster and more complete bone healing in the animal model.

Effect of a freeze-dried CMC/PLGA microsphere matrix of rhBMP-2 on bone healing.

The hypothesis of this research was that implants of poly(lactide-co-glycolide) (PLGA) microspheres loaded with bone morphogenetic protein-2 (rhBMP-2) and distributed in a freeze-dried carboxymethylcellulose (CMC) matrix would produce more new bone than would matrix implants of non-protein-loaded microspheres or matrix implants of only CMC. To test this hypothesis it was necessary to fashion microsphere-loaded CMC implants that were simple to insert, fit precisely into a defect, and would not elicit swelling. Microspheres were produced via a water-in-oil-in-water double-emulsion system and were loaded with rhBMP-2 by soaking them in a buffered solution of the protein at a concentration of 5.4 mg protein per gram of PLGA. Following recovery of the loaded microspheres by lyophilization, matrices for implantation were prepared by lyophilizing a suspension of the microspheres in 2% CMC in flat-bottom tissue culture plates. Similar matrices were made with 2% CMC and with 2% CMC containing blank microspheres. A full-thickness calvarial defect model in New Zealand white rabbits was used to assess bone growth. Implants fit the defect well, allowing for direct application. Six weeks postsurgery, defects were collected and processed for undecalcified histology. In vitro, 60% of the loaded rhBMP-2 released from devices or microspheres in 5 to 7 days, with the unembedded microspheres releasing faster than those embedded in CMC. In vivo, the rhBMP-2 microspheres greatly enhanced bone healing, whereas nonloaded PLGA microspheres in the CMC implants had little effect. The results showed that a lyophilized device of rhBMP-2/PLGA microspheres in CMC was an effective implantable protein-delivery system for use in bone repair.

Skin contraction with pulsed CO2 and erbium:YAG laser.

The purpose of this study was to assess the physical response of skin to laser resurfacing in a real-time, quantitative fashion. The study was designed to assess skin contraction from two opposite standpoints. First, change in tension was measured during laser application while samples were held at constant length. Second, change in length of a sample under no tension was measured during laser treatment. These two disparate analyses represent the two possible extremes of the clinical situation in which skin exists under some tension with some laxity to allow for decrease in length. A custom apparatus with digital interface for skin tension measurements was used to produce single sample tracings of change in skin tension with laser treatment. Length change was measured for individual samples by continuous sonomicrometer readings. Individual sample data were then plotted in a time versus tension/length graph. Skin contracts immediately to a peak level and then relaxes to a sustained plateau level for both CO2 and erbium:YAG lasers. Increased contraction was noted when the beam penetrated into the dermis. Greater peak and plateau contraction is observed after the beam has penetrated into the dermis. Skin contraction varies directly with energy for CO2 and erbium:YAG laser. Findings were similar when skin tension was measured with the sample held at constant length and when length change was measured with the sample under no tension. Char left on the skin after a pass with CO2 laser substantially decreases skin contraction. High-density settings with CO2 laser yield pulse stacking, which effectively irradiates the same portion of tissue with char on it. Skin contraction varies inversely with computer pattern density settings for CO2 laser due to this pulse stacking effect. Density has little effect on skin contraction for the erbium:YAG laser because little char is generated. Histologic analysis identified a zone of coagulated dermis that correlates linearly with skin contraction.

Two lyophilized polymer matrix recombinant human bone morphogenetic protein-2 carriers in rabbit calvarial defects.

We have developed a lyophilized bone morphogenetic protein (BMP) delivery device that can be formulated to control release over 2 to 8 weeks. Bioerodible poly (d,l lactide-co-glycolide) particles loaded with 90 micrograms recombinant human BMP-2 were suspended in either carboxymethylcellulose (CMC) or methylcellulose (MC) implants. Plain CMC and MC implants served as controls, as did a nonimplanted group. A total of 40 rabbits was evaluated histologically 2, 4, or 8 weeks after receiving circular full-thickness 15-mm calvarial defects. MC appeared to prevent prolapse of periosteum and dura into the defects and did not elicit bone growth. Addition of BMP improved the result. CMC implants appeared to encourage bone growth even in the absence of BMP. When BMP was added, new bone formed earlier. CMC may influence new bone formation because it is hydrophilic. MC is less hydrophilic and may cause undue inflammation. Either can be combined with BMP to produce unitary devices that are easy to make and use.

Customized composite reconstruction of extensive midfacial defects.

We describe the treatment of 2 patients with extensive midfacial defects by using a customized composite flap. Autogenous bone was transferred from the iliac crest, revascularized by the rectus muscle, and then transferred as a complete unit by microvascular technique to provide a vascularized bone graft to this area of difficult reconstruction. The bone was shaped and customized to fit the osseous defect. Although the rectus muscle was relatively bulky at the time of harvest, its hearty blood supply permitted trimming and contouring to fit the gap. This technique was based on the principles proposed previously by other authors regarding the vascularization of a free bone graft and its transfer as a composite flap. This technique requires more research and refinement, however this clinical demonstration shows the feasibility of revascularizing a composite flap and the extent to which this technique can be performed.

Critical assessment of the head and neck clinical skills of general surgery residents.

Head and neck surgery is an important part of general surgery. There is, however, little information about the quality of residents' clinical skills in this important field. In an Objective Structured Clinical Examination (OSCE), residents encounter multiple patients with various clinical problems and are rated by faculty members using objective criteria. This study was undertaken to assess the head and neck surgery skills of a group of general surgical residents. Fifty-one general surgery residents examined the same nine patients with head and neck disease. Faculty members graded each clinical interaction according to preset objective criteria. Both actual (e.g., thyroid nodule, oral cancer follow-up examination) and simulated (e.g., dysphagia) patients were used in the OSCE. The reliability of the examination was assessed by coefficient alpha. The construct validity was determined by a two-way analysis of variance with one repeated measure. The reliability was 0.75 for the clinical examination. Performance varied by level of training: Residents performed at a higher level than interns (p < 0.0001), but overall scores were poor (mean score 55%). Important deficits in skills were identified at all levels of training. It is concluded that more attention should be focused on specific outcome assessments of surgical training programs and on strategies for upgrading the clinical skills of surgical residents.

Supraorbital roof fractures: a formidable entity with which to contend.

Supraorbital roof fractures are uncommon. The incidence has been quoted at between 1 and 5%. We believed that the incidence and morbidity of supraorbital roof fractures were actually higher. The charts of 621 inpatients with facial fractures between September 1993 and September 1995 were retrospectively reviewed. Parameters included patient characteristics; mechanism of injury; fracture characteristics; associated clinical, computed tomography, and operative findings; presence of associated skull and frontal sinus fractures; and complication rates. Fifty-eight patients (9.3%) had supraorbital roof fractures, making this one of the largest series. The average age was 31 years and the predominant mechanism of injury was motor vehicle accidents. Sixty-nine percent of the patients had associated skull fractures and 54% had frontal sinus fractures. Thirty-one of the cases were open and 32 cases were treated operatively. Dural tears were present in 14 patients, traumatic encephalocele in 3, proptosis in 6, pulsatile proptosis in 3, orbital apex syndrome in 1, persistent cerebrospinal fluid leak in 3, and meningitis in 5. A majority of the patients had associated intracranial bleeds. The incidence and morbidity of supraorbital roof fractures are much higher than previously thought. The treatment of cases should be individualized, taking into account displacement, association with skull and frontal sinus fractures, dural tears, and intracranial hemorrhage.

High-energy gunshot wounds to the face.

Thirty-three patients with high-energy gunshot wounds to the face were treated at the University of Kentucky Chandler Medical Center between 1976 and 1993. Wounds were classified according to the mass and velocity of the projectile, and the range from weapon to target. More than half the injuries involved multiple facial regions. Twenty patients underwent immediate definitive reconstructive procedures. Intervals between injury and initial nondefinitive reconstruction for the other patients ranged from 1 day to 1 month. Toward the end of the study period, reconstruction was undertaken earlier and more aggressively, and included more attention to primary bone grafting and free tissue transfer. These patients developed fewer problems with infection, long-term scarring, and contracture, and they required fewer operative procedures. There was no operative mortality and none of the patients with self-inflicted injuries reattempted suicide. We conclude that early aggressive treatment of these wounds can produce better structural, functional, and rehabilitative results.

Anatomical study of the blood supply to the skin in rhytidectomy.

This study was undertaken to evaluate the blood supply to the facelift flap by identifying the perforating arteries in the anterior facial region, which is preserved during the undermining of the flap. We have called this medial zone the pedicle area of the flap. Eleven pairs of musculocutaneous perforator arteries supplying the facelift flap were identified, all of them emerging from three main arterial trunks: the facial, the superficial temporal, and the ophthalmic arteries. A rich anastomotic network connected all the vessels; however, we were able to group this network into five basic forms including the ipsilateral and contralateral external and internal carotid arteries. We found these anastomotic links to be constant. We propose that this anatomical feature of the face provides for adequate blood supply even when certain regions have been altered by previous dissection, injury, or congenital deformity. This study confirms previous anatomical findings but also adds specific information regarding the blood supply to the facelift flap that will allow this procedure to be carried out with maximum safety and effectiveness.

The ramifications of immediate reconstruction in the management of breast cancer.

A retrospective review of 50 patients who underwent immediate postmastectomy breast reconstruction was performed to determine the effect of reconstruction on the treatment of these patients. The overall complication rate was 50% (25 of 50). Smoking statistically correlated with an increased rate of wound complications (0 = 0.0001). Obese patients had nearly twice as many wound complications; however, this finding was not statistically significant (P = 0.261). Eleven of the 50 patients underwent reconstruction with a prosthesis, with an overall complication rate of 64% (seven of 11) and a 55% (six of 11) rate of prosthesis loss. Thirty-nine of the 50 patients underwent reconstruction with autologous tissue. Forty-six per cent (18 of 39) of the patients in the autologous group developed complications, and eight required emergent reoperation to prevent flap loss. The wound complication rate was significantly lower for bipedicled transverse rectus abdominis muscle (TRAM) flap reconstructions than for other forms of autologous reconstruction (P = 0.040). total operative time (including mastectomy) was 3.11 hours (range, 2-4 hours) for the prosthetic group and 9.4 hours (range, 5-15 hours) for the autologous group. All but two patients undergoing autologous tissue reconstruction required blood transfusions; an average of 2.4 U of blood was transfused per patient (range, 0-7 U). Only two patients in the prosthetic group required a transfusion. The average hospital stay was 5 days (range, 2-7 days) for the prosthetic group and 8.3 days (range, 5-19 days) for the autologous tissue group.(ABSTRACT TRUNCATED AT 250 WORDS)

Vulvar and vaginal reconstruction after surgical treatment for gynecologic cancer.

The goal of reconstructive surgery is to restore anatomy and function. Sexual activity is an important criterion for reconstruction. Wound healing, postoperative rehabilitation, and a positive self-image also can be restored with the creation of a neovagina or neovulva.

Use of the tram and latissimus dorsi flaps in autogenous breast reconstruction.

Breast cancer has reached near epidemic proportions in this country. It is a devastating blow to the patient and her family. Refinements in breast reconstruction have aided greatly in alleviating this harsh attack on the patient's well being and femininity. Many modalities have been used in the past to achieve breast reconstruction but the use of autogenous tissue and immediate reconstruction have permitted the plastic surgeon the opportunity to regain the normal shape of the breast and at times even obtain an improved aesthetic result. We are still not able to obtain a reconstructed breast that has normal sensation and erogenous stimulation. Patients, however, are demanding more and more of their surgeons as they try to better adapt to their conditions. It is hoped that with the advent of new health care reform these patients will not be deprived of the meaningful advances that have been made to date in this field. To paraphrase a saying attributed to Tagliacozzi: "Our objective is not merely to stamp out disease but it is also to buoy the spirit of the afflicted."

Flap coverage of a large defect after excision of a massive dermatofibrosarcoma protuberans.

We report the case of a 62-year-old man with a huge (25 X 15 cm) dermatofibrosarcoma protuberans of the left flank and abdomen. This is one of the largest dermatofibrosarcoma protuberans lesions reported in the literature. Wide surgical excision of the tumor with 4-cm tumor-free margins created an extensive wound. Wound coverage was performed by transposing an extended rectus abdominis myocutaneous pedicled flap to the defect. Follow-up at 2 years revealed no evidence of local or regional recurrence. We present a brief review of dermatofibrosarcoma protuberans and a discussion of the reconstruction of such large defects.

The cleft earlobe: a review of methods of treatment.

The plastic surgeon must frequently deal with the aesthetic repair of cleft earlobe deformities. Many techniques have been described, each claiming to achieve the best results. In this review of the literature, we discuss the various operations for the treatment of this condition. A classification of the traumatic earlobe clefts is proposed, and some basic principles for surgical treatment are suggested.

Mediastinal dissection and reconstruction for recurrent Hürthle cell carcinoma of the thyroid.

A 69-year-old man presented with recurrent Hürthle cell carcinoma of the thyroid. Successful surgical resection of the recurrent tumor included the larynx and trachea; skin, soft tissues, and lymph nodes of the neck; and the tissues of the anterior mediastinum, including upper sternum and clavicular heads. Coverage of the wound was accomplished with an inferiorly based parasternal fasciocutaneous flap and a long pectoralis myocutaneous flap. This represents the first reported case of Hürthle cell carcinoma treated with mediastinal dissection for extensive local disease. Radical surgical resection should be considered for any patient with Hürthle cell carcinoma of the thyroid in view of the ineffectiveness of nonsurgical modalities.

Sequentially linked free flaps in head and neck reconstruction.

Free tissue transfer has become a useful technique for reconstructing complex three-dimensional defects following the extirpation of head and neck malignancies. This technique, however, may be limited by inadequate vascularity in the recipient site when a patient has been subjected to operative procedures or radiation therapy. The use of serial flaps, with the first flap connected sequentially to the second, reduces the need for dual recipient vessels in a surgical field when two flaps are required for reconstruction. Although this procedure is more technically demanding than more traditional approaches, it can provide a satisfactory, reliable one-stage composite reconstruction under these difficult circumstances. Results have been functionally and aesthetically superior to those achieved with pedicled one-flap reconstructive methods.