Screening of mutations in the PHF8 gene and identification of a novel mutation in a Finnish family with XLMR and cleft lip/cleft palate.

We investigated the prevalence of mutations in the PHD finger protein 8 (PHF8) gene in X-linked mental retardation (XLMR) and facial cleft starting from the original cohort of 7712 patients operated on since 1 January 1950 for cleft lip/cleft palate in the Cleft Centre at the Helsinki University Hospital. From this nationwide material, 18 patients including one family with two male patients with cleft lip/cleft palate and unknown cause of mental retardation (MR) were sequenced for the coding regions and splice sites of the PHF8 gene. A novel missense mutation c.836C>T of the PHF8 gene was identified in a Finnish family with multiple-affected male patients. The mutation resides in exon 8 and changes phenylalanine to serine (F279S) in the functionally important Jmonji C domain of the protein. The clinical phenotype of the male patients was characterized by mild MR, mild dysmorphic features, unilateral cleft lip and cleft palate in one and bilateral cleft lip and cleft palate in the other sibling. The mutation was not present in 200 anonymous blood donors (approximately 300 X-chromosomes). To our knowledge, F279S is the third mutation of the PHF8 gene identified so far.

Endoscopic forehead lift for ptosis of the brow caused by facial paralysis.

Twelve patients with ptosis of the brow caused by facial nerve paralysis were treated by endoscopic forehead lift. Four had an isolated paralysis of the temporal branch of the facial nerve and in eight the entire facial nerve was affected. In two of these patients the facial paralysis was bilateral, caused by Finnish hereditary amyloidosis. A standard subperiosteal endoforehead approach was used and the forehead was fixed by biodegradable pins (n = 8), anchor screws (n = 3), or a suture to the posterior scalp (n = 1). The procedure was a success in 10 patients, and failed in two after initially good results. The endoscopic brow lift is a good method of treating ptosis of the brow caused by paralysis of the temporal branch of the facial nerve.

Mapping of the second locus for the Van der Woude syndrome to chromosome 1p34.

The Van der Woude syndrome (VWS) is a dominantly inherited developmental disorder characterized by pits and/or sinuses of the lower lip, cleft lip and/or cleft palate. It is the most common cleft syndrome. VWS has shown remarkable genetic homogeneity in all populations, and so far, all families reported have been linked to 1q32-q41. A large Finnish pedigree with VWS was recently found to be unlinked to 1q32-q41. In order to map the disease locus in this family, a genome wide linkage scan was performed. A maximum lod score of 3.18 was obtained with the marker D1S2797, thus assigning the disease locus to chromosomal region 1p34. By analyses of meiotic recombinants an approximately 30 cM region of shared haplotypes was identified. The results confirm the heterogeneity of the VWS syndrome, and they place the second disease locus in 1p34. This finding has a special interest because the phenotype in VWS closely resembles the phenotype in non-syndromic forms of cleft lip and palate.

Piperazinylalkyl prodrugs of naproxen improve in vitro skin permeation.

Novel morpholinyl (4a) and piperazinylalkyl (4b-e) esters were synthesized and evaluated in vitro for their properties as bioreversible topically administered dermal prodrugs of naproxen. These ionizable prodrugs exhibited various aqueous solubilities and lipophilicities, depending on the pH of medium. As indicated by octanol-buffer partition coefficients (logP(app)) at pH 7.4, all of the prodrugs were significantly more lipophilic (logP(app)=0.7-3.9) than naproxen (logP(app)=0.3). Furthermore, the most aqueous of the soluble prodrugs (4b-d) were only 2-3-fold less soluble in an aqueous buffer of pH 7.4 ( approximately 30-50 mM) than was naproxen ( approximately 100 mM). At a pH of 5.0, prodrugs showed a generally higher aqueous solubility and similar logP(app) values, compared to naproxen. The chemical and enzymatic hydrolysis of prodrugs at 37 degrees C was investigated in aqueous buffer solutions (pH 5.0 and 7.4) and in 80% human serum (pH 7.4), respectively. The prodrugs showed moderate chemical stability (t(1/2)=15-150 days at pH 5.0), and they were hydrolyzed enzymatically to naproxen, with half-lives ranging from 0.4 to 77 min. In permeation studies using post-mortem human skin in vitro, the flux of naproxen was 6.5 and 1.6 nmol/cm(2). h in a saturated aqueous buffer vehicle of pH 7.4 and 5.0, respectively. Among the prodrugs, two piperazinyl derivatives (4c and 4d) resulted in a 9- and 4-fold enhancement of permeation, respectively, when compared to naproxen itself at pH 7.4. 4c also resulted in a significantly (4-fold) better permeation than naproxen at pH 5.0. In conclusion, piperazinyl esters improved skin permeation of naproxen and are promising prodrugs of naproxen for topical drug delivery.

Synthesis and in vitro evaluation of novel morpholinyl- and methylpiperazinylacyloxyalkyl prodrugs of 2-(6-methoxy-2-naphthyl)propionic acid (Naproxen) for topical drug delivery.

Various novel morpholinyl- (3a,b) and methylpiperazinylacyloxyalkyl (3c-f) esters of 2-(6-methoxy-2-naphthyl)propionic acid were synthesized and evaluated in vitro for topical drug delivery as potential prodrugs of naproxen (1). Compounds 3a-f were prepared by coupling the corresponding naproxen hydroxyalkyl ester with the morpholinyl- or (4-methyl-1-piperazinyl)acyl acid in the presence of dicyclohexylcarbodiimide (DCC) and 4-(dimethylamino)pyridine (DMAP) and quantitatively hydrolyzed (t(1/2) = 1-26 min) to naproxen in human serum. Compounds 3c-f showed higher aqueous solubility and similar lipophilicity, determined by their octanol-buffer partition coefficients (log P(app)), at pH 5.0 when compared to naproxen. At pH 7.4 they were significantly more lipophilic than naproxen. The best prodrug 3c led to a 4- and 1.5-fold enhancement of skin permeation when compared to naproxen at pH 7.4 and 5.0, respectively. The present study indicates using a methylpiperazinyl group yields prodrugs that are partially un-ionized under neutral and slightly acidic conditions, and thus, a desirable combination is achieved in terms of aqueous solubility and lipophilicity. Moreover, the resulting combination of biphasic solubility and fast enzymatic hydrolysis of the methylpiperazinylacyloxyalkyl derivatives gave improved topical delivery of naproxen.

Synthesis and in vitro evaluation of aminoacyloxyalkyl esters of 2-(6-methoxy-2-naphthyl)propionic acid as novel naproxen prodrugs for dermal drug delivery.

PURPOSE: To synthesize and evaluate various novel aminoacyloxyalkyl esters of naproxen (3a-i) and naproxenoxyalkyl diesters of glutamic and aspartic acids (3j-m) as potential dermal prodrugs of naproxen. METHODS: The prodrugs 3a-m were synthesized, and their aqueous solubilities, lipophilicities and hydrolysis rates were determined in a buffered solution and in human serum. The permeation of selected prodrugs across excised postmortem human skin was studied in vitro. RESULTS: The aminoacyloxyalkyl prodrugs showed higher aqueous solubilities and similar lipid solubilities, in terms of octanol-buffer partition coefficients (log Papp) at pH 5.0, when compared with naproxen. At pH 7.4 the prodrugs were significantly more lipophilic than naproxen. Prodrugs 3a-i showed moderate chemical stability in aqueous solutions at pH 5.0 and were rapidly converted to naproxen in human serum (t1/2 = 4-19 min). The selected aminoacyloxyalkyl prodrugs possessed a higher flux across the skin than naproxen, with a maximum enhancement of 3-fold compared to naproxen. Prodrugs 3j-m showed poor aqueous solubility and permeation across the skin. CONCLUSIONS: Combinations of adequate aqueous solubility and lipophilicity of naproxen aminoacyloxyalkyl prodrugs having fast rates of enzymatic hydrolysis resulted in improved dermal delivery of naproxen.

In vitro evaluation of acyloxyalkyl esters as dermal prodrugs of ketoprofen and naproxen.

A series of acyloxyalkyl esters of ketoprofen and naproxen were synthesized and investigated as topical prodrugs with the aim of improving the dermal delivery of the drugs. In addition, some hydroxyalkyl esters of ketoprofen and naproxen were synthesized as possible intermediates of acyloxyalkyl prodrugs. All of the prodrugs were more lipophilic than their parent molecules, as evaluated by drug partitioning between 1-octanol and phosphate buffer at pH 7.4 (log Papp). However, their solubilities in aqueous solutions decreased markedly compared with the parent molecules. The prodrugs were stable toward chemical hydrolysis in aqueous solutions (pH 7.4), but were hydrolyzed to the parent drug both in 80% human serum and in human skin homogenate, with half-lives ranging from 4 to 137 min and from 13 to 403 min, respectively. The abilities of the selected naproxen acyloxyalkyl prodrugs to deliver naproxen through excised human skin were evaluated. Generally, the prodrugs showed similar dermal delivery as the parent drug through cadaver skin. In the present series of lipophilic prodrugs of naproxen, the prodrug with the highest aqueous solubility was the most effective prodrug to deliver naproxen through the skin.

Gait patterns after free flap reconstruction of the foot sole.

Walking patterns after reconstruction of soft tissue defects of the sole of the foot with free flap were studied in seven patients. Measurements included walking technique, ground reaction forces and electromyographic (EMG) activity during the gait cycle. The results showed that only one patient walked symmetrically; the remaining six had many differences between the legs, the asymmetry usually being noticed when the subjects walked barefoot. The most common change was the shortening of the stance phase of the injured foot. The maximal vertical ground reaction force decreased in the braking phase and in the push-off phase. Quantitative differences in EMG between the legs were less than expected. The present study gives some support to the hypothesis that altering the pattern of gait may contribute to soft tissue stability after resurfacing of the sole.

[Covering the foot sole and heel with pedicled and free transplants]. / Deckung der Fusssohle und Ferse mit gestielten und freien Transplantaten.

Thirty patients suffering from defects of the weight-bearing areas of the sole and heel were analyzed employing various methods. Sensibility was analyzed quantitatively. The level of sensibility of the flaps was found to be variable, but it had no effect on soft-tissue stability. Sensory recovery in the flaps was slow and unpredictable. Gait analysis showed that patients supplied with footwear walked relatively symmetrically even though there was a tendency to protect the flap by decreased weight-bearing. Postural control as analyzed by a sway plate was impaired only in patients with flaps having very poor sensibility. Thin skin flaps were found to be most suitable for degloving defects and muscle flaps with split-skin coverage for deep defects. The scapular flap was often too thick and was prone to develop superficial ulcerations on the weight-bearing areas.

Resurfacing and sensory recovery of the sole.

(1) There is very little quantitative objective information on sensory recovery after resurfacing of the sole. However, based on results obtained by quantitative rather than qualitative methods to measure sensibility, there is little reason to expect a high level of sensory recovery, at least in free flaps. There is no evidence to prove or refute that by using neurosensory flaps or other forms of neurotization, a higher level of sensibility can be achieved, and therefore no reason why such a flap should be specifically selected when reconstructing the sole. Also, the sensibility of the flap has not been shown to be related to the soft-tissue stability after resurfacing of traumatic defects. (2) There are no firm guidelines to flap selection. If possible, a local flap should be selected. If a free-tissue transfer is necessary, a safe choice is for the surgeon to select a familiar flap with a long pedicle of sufficient caliber. The flap should be well tailored into the defect, and excessively thick flaps should be avoided. If in doubt, the donor area can be measured with ultrasonography and compared with the depth of the defect. Muscle flaps should be used to cover deep, irregular, or infected defects, whereas skin flaps are suitable for resurfacing superficial degloving tissue loss. (3) The most important reason for recurrent breakdowns after reconstruction may be abnormally high pressure points in the sole. Bony deformities should be identified preoperatively and removed during the operation. Patient education in meticulous foot care and custom-made insoles or footwear should be provided to prevent reulcerations. (4) The ideal method of resurfacing of the sole would also provide normal sensibility to the skin. With free-flap transfer this cannot be achieved with current techniques. Before new methods of neural reconstruction can be recommended for general use, their efficacy in improving sensibility should be documented by prospective studies using quantitative methods of measurement. If a reduction of breakdowns is claimed, biomechanical studies should be employed to exclude this being merely due to elimination of abnormal pressure points in the sole.

Suitability of the scapular flap for reconstructions of the foot.

Eighteen patients with mainly a traumatic soft-tissue defect of the foot underwent reconstruction with a microvascular free scapular flap. Of the 17 successful transfers, 13 were to the weight-bearing parts of the foot. The stability and contour of the flaps were assessed after an average follow-up time of 3 years (range 1 to 5 years). The thicknesses of the scapular donor site and flap and the recipient site were measured by an ultrasound technique. The resistance of the flap to shear was measured with a dynamometer. The ultrasound measurements aided in refining our operative technique. In early cases, the flap thickness after transfer could be more than double what it was in the donor area. With proper tightening, the thickness could be reduced, with improvement in contour but no increase in soft-tissue stability or shear resistance of the flap. Without proper tightening, the scapular flap tended to be redundant when transferred to the foot. For good results, the patient should be lean, since the optimal thickness of the scapular donor site was less than 6 mm and the maximum thickness should not exceed 8 to 10 mm. The differences in shear resistance between the flaps were not associated with the soft-tissue stability of the reconstruction. The relative laxity of the flap on the plantar surface was found by several patients to be subjectively unpleasant. Although good contour could be achieved when covering the plantar heel, the tendency of the flap to develop abrasions and superficial breakdowns made it unsatisfactory for covering this area.(ABSTRACT TRUNCATED AT 250 WORDS)

Patterns of recovery of sensibility in free flaps transferred to the foot: a prospective study.

In a prospective study, the recovery of mechanical sensibility in five free flaps transferred to the foot was assessed qualitatively and measured with a quantitative method. The detection thresholds for 20, 80, and 240 Hz vibration frequencies were determined preoperatively from the recipient area and postoperatively from the flap at intervals of two to six months up to 14 to 21 months. Detection thresholds to mechanical vibration could be measured in the flaps immediately after operation before any sensory innervation could have taken place. The immediate postoperative thresholds were similar to the preoperative values obtained from an existing full-thickness defect; and where surgery had created a considerably deeper defect, they were elevated. The recovery of mechanical sensibility in the transferred flaps was found to be slow and incomplete. In three patients, after one year of follow-up, the flaps showed qualitative improvement in sensibility accompanied by a moderate lowering of detection thresholds obtained with 240 Hz vibration frequency. In two other flaps, there was no improvement in sensibility. In the reconstructions that had good soft-tissue stability, the flap was resistant to breakdown even before sensory recovery.

[Reconstruction of the foot using free flaps]. / Fussrekonstruktionen mit freien Lappen.

Microvascular free-flap reconstruction of the foot was performed during a seven-year period in 25 patients to repair soft tissue loss caused mainly by traumatic injury. A scapular flap was used in fifteen cases, a dorsalis pedis in three, a radial forearm in two, a latissimus dorsi in two, and a gluteal thigh, rectus abdominis and tensor fascia lata each in one case. Two flaps were lost because of postoperative vascular complications. Soft-tissue stability was excellent or good in all of the seven patients with a successful free-flap transfer to the non-weightbearing part of the foot and in eleven of the sixteen patients with a flap on the weightbearing part of the foot. Four patients had frequent superficial ulcerations of the graft but only one flap was completely unstable. Sixteen patients had normal or near normal ability to walk. Significant gait problems in seven patients were caused by skeletal deformity of the reconstructed foot or other associated injuries of the lower limb rather than the reconstruction itself. The best fit and contour was provided by thin skin flaps such as the dorsalis pedis and radial forearm flaps while the scapular flap offered a more inconspicuous donor site and could be used to cover large defects. Sensibility of the flaps assessed by clinical methods was less than normal in all cases. Good sensibility was found in the flaps of two children, fair in six, poor or very poor in fifteen, and two flaps had no sensibility. The level of sensibility was not related to the soft-tissue stability of the flap.(ABSTRACT TRUNCATED AT 250 WORDS)

Mechanical sensibility in free and island flaps of the foot.

Mechanical sensibility in 20 free skin flaps and four dorsalis pedis island flaps, used for the reconstruction of foot defects, was analyzed with conventional clinical methods and by determining sensibility thresholds to vibration frequencies of 20, 80, and 240 Hz. To eliminate inter-individual variability, a score was calculated for each frequency by dividing the thresholds determined for each flap with values obtained from the corresponding area on the uninjured foot. The soft tissue stability of the reconstruction was assessed. Patients were divided into three groups according to the scores. In the group of flaps with the best sensibility, the threshold increases were low at all frequencies. In the group with intermediate sensibility, the relative threshold increases were greater, the higher the frequency. In the group with the poorest sensibility, no thresholds were obtained with 240 Hz frequency and the thresholds increases were very high at all frequencies. Sensibility was not related to the length of follow-up time, nor to the type or size of the flap. However, flap sensibility was closely associated with that of the recipient area, where sensibility was usually inferior to that of normal skin. The island flaps generally had better sensibility than the free flaps. There was a good correspondence between the levels of sensibility determined by clinical and quantitative methods. The quantitative data on the level of sensibility obtained with the psychophysical method were found to be reliable and free from observer bias, and are therefore recommended for future studies. The degree of sensibility may have contributed to, but was not essential for, good soft-tissue stability of the reconstruction.

Mechanical sensibility of the sole of the foot determined with vibratory stimuli of varying frequency.

The mechanoreceptive properties of the sole of the foot were determined by measuring the detection thresholds to vibratory stimuli of 20, 80, and 240 Hz frequency and 300 ms duration. The thresholds were measured at six different sites on the left sole and at toes 1 and 3 with probes of 2 and 8 mm diameter connected to the moving coil of an electromechanical vibrator. The subject sat in an armchair during the experiments, with the left leg supported horizontally by a vacuum cast positioned on a table. Six subjects participated in the experiments. A simple method of limits was used to make the measurements. Lower average thresholds were obtained with higher vibration frequencies, the average thresholds varying between 40-90 microns at 20 Hz and well below 10 microns at 240 Hz. The major decrease in thresholds occurred between 20 and 80 Hz. Interindividual variability in thresholds was large, but the threshold curves obtained from different subjects and from different stimulation points were of the same general shape. The highest thresholds were measured from the toes, but this regional differences in sensibility was obtained only at the higher vibration frequencies. Comparison of the threshold values at the sole with those found with similar stimuli at the thenar eminence and middle fingertip indicates that the mechanoreceptor mechanisms transmitting information about low-frequency vibration in the sole are similar to those in the palmar skin of the hand.