Cryopreservation of Kalopanax septemlobus embryogenic callus using vitrification and droplet-vitrification.

A cryopreservation protocol has been developed for embryogenic callus cultures of castor aralia (Kalopanax septemlobus), a deciduous tree which is widely used in oriental medicine and in landscape design. Three preculture treatments, four loading and six vitrification solutions were tested in a vitrification procedure. Preculture of embryogenic callus (EC) with high sucrose concentrations (up to 0.7 M) showed no effect on regrowth after cryopreservation. Loading for 20 min at ambient temperature improved regrowth of cryopreserved EC by 70-75 percent compared with non-loaded samples, regardless of the composition of the loading solution. Among vitrification solutions, the highest regrowth of 95-100 percent after cryopreservation was obtained after incubation of EC in a vitrification solution A3-80 percent comprising (w/v) 33.3 percent glycerol + 13.3 percent DMSO + 13.3 percent EG + 20.1 percent sucrose for 40 min at 0°C. Profiling of crystallization and recrystallization events using differential scanning calorimetry (DSC) confirmed that freezing injury was minimized in samples after loading and cryoprotection with this vitrification solution. Unlike many other papers, the droplet-vitrification protocol did not produce higher post-cryopreservation regrowth of Kalopanax EC, compared with the vitrification procedure. When samples are sufficiently cryoprotected during VS treatment, vitrification using cryovials may be preferred, since droplet-vitrification is more complex and requires skilled personnel. Cryopreserved callus grew rapidly and produced numerous somatic embryos, which developed similarly to embryos obtained from non-cryopreserved samples.

Comparison of medial and lateral meniscal transplantation with regard to extrusion of the allograft, and its correlation with clinical outcome.

We compared extrusion of the allograft after medial and lateral meniscal allograft transplantation and examined the correlation between the extent of extrusion and the clinical outcome. A total of 73 lateral and 26 medial meniscus allografts were evaluated by MRI at a mean of 32 months (24 to 59) in 99 patients (67 men, 32 women) with a mean age of 35 years (21 to 52). The absolute values and the proportional widths of extruded menisci as a percentage were measured in coronal images that showed maximum extrusion. Functional assessments were performed using Lysholm scores. The mean extrusion was 4.7 mm (1.8 to 7.7) for lateral menisci and 2.9 mm (1.2 to 6.5) for medial menisci (p < 0.001), and the mean percentage extrusions were 52.0% (23.8% to 81.8%) and 31.2% (11.6% to 63.4%), respectively (p < 0.001). Mean Lysholm scores increased significantly from 49.0 (10 to 83) pre-operatively to 86.6 (33 to 99) at final follow-up for lateral menisci (p = 0.001) and from 50.9 (15 to 88) to 88.3 (32 to 100) for medial menisci (p < 0.001). The final mean Lysholm scores were similar in the two groups (p = 0.312). Furthermore, Lysholm scores were not found to be correlated with degree of extrusion (p = 0.242). Thus, transplanted lateral menisci extrude more significantly than transplanted medial menisci. However, the clinical outcome after meniscal transplantation was not found to be adversely affected by extrusion of the allograft.

Congenital meniscoid articular disc of the triangular fibrocartilage complex.

We report the case of a 24-year-old man with a congenital meniscoid articular disc of the triangular fibrocartilage complex with extensor carpi ulnaris tenosynovitis. His young age, the normal articular cartilage, the lack of degenerative changes at the margins of the defect and its bilateral occurrence made this diagnosis likely. A congenital defect of the articular disc of the triangular fibrocartilage complex should not be misinterpreted as a traumatic rupture and is usually asymptomatic.

Effects of dietary arginine supplementation during gestation and lactation on the performance of lactating primiparous sows and nursing piglets.

A 2 x 2 factorial arrangement of treatments in a randomized block design was used to determine the effects of dietary Arg supplementation during gestation and lactation on the lactation performance of 38 first-parity sows. At 30 d of gestation, pregnant gilts were allotted based on BW to 1 of 2 diets supplemented with 1% L-Arg.HCl or 1.7% L-Ala (isonitrogenous control). After farrowing, sows were further allotted based on BW within previous gestation treatment groups to 1 of 2 lactation diets supplemented with 1% L-Arg.HCl or 1.7% L-Ala (isonitrogenous control). All gestation diets contained 3.1 Mcal/kg and 12.2% CP (as is) and were fed 2 kg/d in 2 equally sized meals, whereas all lactation diets contained 3.2 Mcal/kg and 18.6% CP (as is) and were fed ad libitum. Litter size was standardized to 10 piglets by cross-fostering within 24 h postfarrowing. On a weekly basis, BW and backfat (BF) thickness of sows, as well as piglet BW were measured, and blood and milk samples were obtained from the sows. Number of days from weaning to estrus and ADFI were also recorded. There were no differences in BW, BF thickness, ADFI, or days until return to estrus among treatment groups. There was no effect of the gestation diet or a gestation x lactation diet interaction on any parameter measured. On d 7 of lactation, plasma concentrations of Arg and insulin in sows, as well as concentrations of most AA in milk, were greater (P < 0.05) in response to Arg supplementation during lactation compared with the control. Weight gain of piglets from sows fed the Arg-supplemented diet during lactation was greater between d 0 and 7 (P < 0.01) and between d 0 and 21 (P < 0.05) of lactation compared with piglets from sows fed the control diet. Collectively, results from this study indicate the potential beneficial effects of dietary Arg supplementation in improving the lactation performance of first-parity sows.

Production of eleutherosides in in vitro regenerated embryos and plantlets of Eleutherococcus chiisanensis.

High frequency somatic embryogenesis of Eleutheorcoccus chiisanensis was achieved through suspension culture of embryogenic cells in hormone-free Murashige and Skoog liquid medium supplemented with 30 g sucrose l-1. Cotyledonary somatic embryos were germinated and converted into plantlets using 20 microM: gibberellic acid which were then grown in a 10 l airlift bioreactor. HPLC analysis revealed the accumulation of eleutheroside B, E and E1 in the embryos and plantlets. Thus mass production of embryos and plantlets of E. chiisanensis can be achieved in liquid cultures and the biomass produced may become an alternative source of eleutherosides.

Autologous cardiomyocyte transplantation using a biodegradable polymer scaffold.

BACKGROUND: To achieve a more reliable way of transplanting cardiomyocytes, we conducted an autologous cardiomyocyte transplantation using a biodegradable scaffold, instead of a syringe injection, as a vehicle for transporting cells in an ovine myocardial infarction model. MATERIALS AND METHODS: A myocardial infarction was created in sheep using sequential ligation of the homonymous artery and its diagonal branch. Autologous cardiomyocytes from the right ventricular infundibulum were cultured and seeded onto a biodegradable polymer scaffold. Three months after creating myocardial infarction, the two animals were re-anesthetized and cardiomyocyte-seeded scaffolds were implanted in the infarcted area. The animals were kept alive for a further month, and then sacrificed for postmortem heart examinations. Light microscopic analysis and an immunohistochemical study for myoglobin were performed. RESULTS: On postmortem gross examinations, the polymer scaffolds were visible in the background of well-demarcated thin-walled anteroseptal myocardial infarcts. Microscopic analysis showed abundant myoglobin-stained cells between the fiber strands of the polymer scaffolds. However, there is a possibility that some of these cells might have been giant cells reacting to foreign material. CONCLUSION: The transplantation of cultured autologous cardiomyocytes into an infarct region using a biodegradable scaffold instead of syringe injection provides another promising option for cardiomyocyte transplantation, which warrants further study.

Effect of Ca2+ channel blockers, external Ca2+ and phospholipase A2 inhibitors on t-butylhydroperoxide-induced lipid peroxidation and toxicity in rat liver slices.

OBJECTIVES: This study was undertaken to examine the effect of oxidant on lipid peroxidation and lethal cell injury in rat liver slices. METHODS: t-Butylhydroperoxide (t-BHP) was employed as a model of an oxidant. The lipid peroxidation and lethal cell injury were estimated by measuring the formation of malondialdehyde (MDA) and lactate dehydrogenase (LDH) release, respectively. RESULTS: t-BHP increased lipid peroxidation and LDH release in a dose-dependent manner over concentrations of 0.5-10 mM. t-BHP-induced lipid peroxidation was completely prevented by an antioxidant, N,N-diphenyl-p-phenylenediamine (DPPD), but LDH release was partially decreased. Both t-BHP-induced lipid peroxidation and LDH release were significantly protected by iron chelator, deferoxamine, sulfhydryl reducing agent, dithiothreitol and glutathione. Ca2+ channel blockers, verapamil, diltiazem and nifedipine exerted a significant protective effect against t-BHP-induced lipid peroxidation and LDH release. By contrast, addition of external Ca2+ chelator, ethylene glycol bis(b-aminoethyl ether)-N,N-tetraacetic acid (EGTA) did not alter t-BHP-induced lipid peroxidation, whereas t-BHP-induced lethal cell injury was significantly prevented. Phospholipase A2 (PLA2) inhibitors, mepacrine and butacaine produced a partial protective effect. CONCLUSIONS: These results suggest that t-BHP induces cell injury by lipid peroxidation-dependent and -independent mechanisms which can be partially prevented by Ca2+ channel blockers and PLA2 inhibitors.

Reduction of chronic ciclosporin nephrotoxicity by thromboxane synthase inhibition with OKY-046.

Ciclosporin A (CsA) is a potent immunosuppressive agent which is extremely effective in controlling allograft rejection and in the treatment of autoimmune disease and nephrotic syndrome. Unfortunately, its use is limited by chronic, irreversible nephrotoxicity. Administration of CsA induces renal vasoconstriction, causing a reduction in renal blood flow. An alteration of the prostaglandin-thromboxane cascade may be involved in the vasoconstriction. We studied the role of thromboxane A2 in CsA nephrotoxicity and the ability of a thromboxane synthase inhibitor, OKY-046, to reduce the CsA nephrotoxicity. Daily administration of CsA 25 mg/kg for 28 days to Sprague-Dawley rats resulted in increased excretion of urinary thomboxane B2 (47.9+/-11.5 vs. 27.2+/-9.7 ng/24 h; p<0.05) and decreased creatinine clearance (0.25+/-0.07 vs. 0.43+/-0.17ml/min/100 g; p<0.01) as compared with administration of vehicle only. Histologically, large numbers of lysosomes in the tubular epithelium were characteristic. Coadministration of OKY-046 prevented both the rise in urinary thromboxane B2 excretion (40.0+/-11.8 ng/24 h) and the reduction in the creatinine clearance (0.44+/-0.11 ml/min/100 g). The severity of the histological changes was significantly diminished. Selective inhibition of thromboxane production with OKY-046 may be valuable in the attenuation of CsA nephrotoxicity.

The stimulated gracilis neosphincter for fecal incontinence: a new use for an old concept.

The stimulated gracilis neosphincter is a viable procedure in selected patients with fecal incontinence. The aim of this paper is to review the technique of this staged operative procedure and review the problems and complications. Stage 1 consists of the vascular "delay" of the gracilis muscle and the creation of a temporary stoma. Stage 2 consists of transposition of the muscle around the anus with implantation of the stimulator. Low-frequency electrical stimulation is applied to the muscle for 12 weeks, after which stage 3 (stoma closure) is undertaken. From March of 1993 to March of 1995, 14 patients (9 females and 5 males) with a mean age of 44 years (range 20 to 67 years) underwent the procedure. Two patients died within 1 year of the operation from unrelated causes. Two patients developed anal stenosis and required permanent stomas. Other complications noted during ascent of the learning curve included seroma, excoriation of the skin above the stimulator, transposition of the stimulator, premature battery discharge, wound infection, rupture of the gracilis tendon, fatigue during programming sessions, and electrode displacement or fibrosis from the nerve. However, 8 of the 10 eligible patients had stoma reversal; the manometric results showed an average mean squeeze pressure that increased from 43 mmHg prior to surgery to 151 mmHg after the operation (p < 0.01). Based on an objective functional questionnaire, 60 percent of the patients who could be evaluated reported improvement in continence, social interactions, and the quality of their life. In conclusion, despite a steep learning curve, the stimulated gracilis operation is a viable operation for selected patients with severe incontinence.

Nutrition programs through public health centers in Korea.

In the period of confusion after the Korean War, there was an absolute shortage in the volume of food. The intake of energy and all other nutrients fell short of the recommended dietary intake. To solve the food shortage, there were many food assistant programs through the government channel. With the economic developments during past years, now, there are enough foods for everyone. Thus, food assistant program, which has been the only nutrition program in the government, is no longer needed except for low-income families. Currently, there are not enough nutrition programs in the government. In 1994, about 30 public health center start nutrition programs with the help of Korean Dietetic Association. In this study, kinds of nutrition programs, age, sex and number of participants, the content of programs are studied. The effects of nutrition programs are partially evaluated. Also, problems with current nutrition programs are discussed. Finally, future directions of nutrition programs in the public health centers are discussed.

Stimulated gracilis neosphincter operation. Initial experience, pitfalls, and complications.

PURPOSE: The stimulated gracilis neosphincter is accepted as a viable option in select patients with fecal incontinence. The aim of this study was to review the initial problems and complications. METHODS: A prospective analysis of all patients who underwent this procedure was undertaken. Stage I consisted of the distal vascular delay of the muscle and creation of a temporary stoma. Stage II was the transposition of the muscle and implantation of the stimulator and electrodes. Low frequency electrical stimulation was applied to the muscle for 12 weeks, after which Stage III (stoma closure) was undertaken. RESULTS: From March 1993 to December 1995, 17 patients (9 females and 8 males) with a mean age of 42.2 (range, 19-72) years underwent the procedure. One patient died from pancreatitis and another from small-bowel adenocarcinoma, three and six months after the procedure, respectively. Two patients (one with Crohn's disease) required permanent stomas. One additional patient required a permanent stoma because of lead fibrosis. Other complications noted during ascent of the learning curve included seroma of the thigh incision, excoriation of the skin above the stimulator, fecal impaction, anal fissure, parastomal hernia, rotation of the stimulator, premature battery discharge, fracture of the lead, perineal skin irritation, perineal sepsis, rupture of the tendon, tendon erosion, muscle fatigue during programming sessions, and electrode displacement from the nerve or fibrosis around the nerve. However, ultimately after rectification of these problems, 13 of the 15 eligible patients had stoma reversal. Manometric results showed an average basal pressure of 43 mmHg and an average maximum squeeze pressure that increased from 36 mmHg before surgery to 145 mmHg by stimulation (P < 0.01). Based on objective functional questionnaires, 9 of 15 (60 percent) evaluable patients reported improvement in continence, social interactions, and quality of life. Three of these nine patients require daily use of enemas. CONCLUSION: Although the stimulated gracilis operation is a feasible procedure for selected patients with severe incontinence, the learning curve is steep. Although the ultimate outcome in a selected group of patients can be very gratifying, major technical modifications are required before use beyond a research protocol setting. Furthermore, patients must have the psychological strength, emotional commitment, and financial resources that may be necessary for multiple revisional surgeries or ultimate device failure.

Oromandibular reconstruction using a keel-shaped modification of the radial forearm osteocutaneous flap.

The keel-shaped modification for harvest of the radial forearm osteocutaneous flap has been used to reconstruct 19 oromandibular defects in 18 patients. Fourteen men and 4 women ranging in age from 22 to 72 years have undergone composite mandibular reconstruction, with follow-up ranging from 3 to 36 months. Sixteen patients (17 reconstructions) had resection of advanced malignancies, and 2 patients sustained shotgun wounds. Twelve symphyseal and 7 lateral or posterior defects were reconstructed with donor radius bone ranging in length from 5 to 13.5 cm. Double osteotomies were performed in 7 patients. Two skin paddles were used in 4 patients to provide simultaneous intraoral lining and external skin coverage. The radial forearm osteocutaneous flap is still an excellent choice for oromandibular reconstruction. Anterior and lateral composite mandibular defects were satisfactorily reconstructed both aesthetically and functionally using the keel-shaped modification of the radial forearm flap. Donor-site problems were uncommon and minor, and long-term forearm function was minimally affected. Radius fracture occurred in only 1 patient.

Omentum graft for intractable subdural empyema.

A subdural empyema developed in a young man after craniotomy for evacuation of a hematoma in a sylvian fissure arachnoid cyst and the subdural space. Despite prolonged systemic and subdural antibiotic administration and a debridement of the subdural space, infection persisted, as evidenced by persistent fever, an elevated white blood cell count, and an extremely low cerebrospinal fluid glucose level. The infection was cured after a second debridement operation where microsurgically revascularized free omentum was used to obliterate the cyst and to cover the cerebral hemisphere in the craniotomy defect. The use of vascularized free omentum may prove useful in cases of refractory cranial wound infection and cerebrospinal fluid fistulas.

The vascular anatomy of rectus abdominis musculocutaneous flaps based on the deep superior epigastric system.

Radiographic studies of the deep superior epigastric artery (DSEA) and its connections within the soft tissues of the abdominal wall were performed in 64 fresh cadavers. The patterns of anastomosis between the deep superior epigastric artery and the deep inferior epigastric artery (DIEA) were noted. Type I (29 percent) revealed a single deep superior epigastric artery and deep inferior epigastric artery, type II (57 percent) revealed a double-branched system of each vessel, and type III (14 percent) revealed a system of three or more major branches. In each case, the two systems were united by choke vessels in the segment of muscle above the umbilicus. The supply to the various transverse and vertical skin flaps from the deep superior epigastric artery was defined as a series of captured anatomic territories bounded by choke vessels. The upper transverse and vertical flaps had the best supply, and the TRAM flap had the most tenuous supply. Midline crossover occurs predominantly in the subdermal plexus and on the surface of the rectus sheath. Modifications of the design of the TRAM flap, the case for a delay procedure, the wisdom of including a strip of anterior rectus sheath, and the risks of splitting the muscle with respect to its nerve supply and vascular patterns are discussed on an anatomic basis.

Infections of the cardiac suture line after left ventricular surgery.

PURPOSE: Infections of the cardiac suture line after left ventricular surgery are rare but may be fatal if not diagnosed promptly and treated effectively. In order to alert physicians to this entity, we reviewed data from three patients who presented at the Cleveland Clinic Hospital and from 22 patients in the literature. PATIENTS AND METHODS: The three patients in the current report underwent treatment at the Cleveland Clinic Hospital. Additional cases of infection of the left ventricular suture line were identified by reviewing the English literature pertaining to surgery for left ventricular aneurysms, pseudoaneurysms, and postoperative cardiac infections. RESULTS: Infection presented on average 16 months after surgery with cardiocutaneous fistulae, chest wall masses, hemoptysis or other pleuropulmonary symptoms, or systemic illness with bacteremia resembling endocarditis. Staphylococci and gram-negative bacilli were the most frequent pathogens. Diagnosis was often delayed and mortality was high. Left ventricular false aneurysms were identified in 15 of the 25 patients. Bleeding from sinuses in the chest wall or epigastrium or repeated hemoptysis were important clinical clues. In some instances, ill-advised surgical or instrumental procedures precipitated life-threatening hemorrhage. Treatment with antibiotics alone was insufficient. Excision of all infected sutures and Teflon pledgets and adequate debridement of the infected suture line were required to achieve cures. CONCLUSION: Since infection of the left ventricular suture line has protean clinical manifestations and may present months or years after the initial surgery, a high index of suspicion is of paramount importance in diagnosing the condition. Institution of cardiopulmonary bypass and reoperation through median sternotomy is recommended to achieve a cure.

The relationship of the extrinsic blood supply to regeneration in graft reconstructed peripheral nerves.

Median nerves of rats were reconstructed with conventional or vascular nerve grafts. After 2, 3 and 4 weeks, Allen Video-Enhanced Contrast, Differential Interference Contrast (AVEC-DIC) microscopy revealed axonal transport in most preparations, with varying degrees of myelination. Radio-isotope tracer was measured in the nerve. Two and 4 week measurements revealed no difference between the graft types. At 3 weeks the vascular graft group exhibited transport along the entire length of the nerve in contrast to a relatively abbreviated path length in the conventional graft group. Nerve conduction velocities (NCV) were measured proximal to, within and distal to the grafts. Three week NCV showed no difference between the graft types. The 4 week NCV revealed normal values in the vascular graft group at points distal to and within the graft. Significant slowing was seen in the conventional grafts at both points.

Lumbar hernia after latissimus dorsi flap.

The authors present what they believe to be the first recorded case of a lumbar hernia following latissimus dorsi myocutaneous flap procedure. The authors discuss the diagnosis by physical examination and x-ray confirmation, pointing out the importance of not mistaking the hernia for a seroma. Suggestions for avoiding the problem are given, relating primarily to preservation of the fascia underlying the distal latissimus aponeurosis.