[Ocular manifestation of progressive essential telangiectases]. / Manifestation oculaire de télangiectasies essentielles progressives.

Generalized essential telangiectasia is a rare skin disorder characterized by the development of dilated venules beginning at the lower extremities and progressively spreading out to the rest of the body. It is not related to any known affection and is therefore considered to be essential. The condition tends to affect mostly women in their midthirties. We report the case of a 49-year-old healthy man worried by the progressive extension of dilated vessels on the conjunctivae of both eyes and the progressive worsening at the condition over the last five years. On clinical examination he presented also telangiectasies on both arms and legs which had also grown worse over the last five years. All further investigations and complementary examinations showed no disease. A cutaneous biopsy showed dilated venules in the superior dermis supporting the diagnosis of telangiectases. Several differential diagnoses are discussed. With confirmation of the clinical diagnosis we started treatment with oral tetracycline. Because of the poor response we stopped treatment after a period of three months.

Recombinant expression of alliin lyase from garlic (Allium sativum) in bacteria and yeasts.

Recombinant garlic alliin lyase was produced in Escherichia coli, Saccharomyces cerevisiae, and Pichia pastoris. A cDNA clone was obtained from garlic bulbs by PCR and introduced into suitable bacterial and yeast expression vectors. The recombinant alliin lyase forms inclusion bodies in all three host organisms, which are deposited in the cytoplasm. After cell lysis and harvesting by centrifugation, the inclusion bodies were solubilized in Zwittergent 3-14 solution and refolded by stepwise dilution. Specific alliin lyase activity could be recovered by this procedure.

[Neurophysiologic follow-up in patients with hypophyseal adenoma operations]. / Neurophysiologische Verlaufskontrollen bei Patienten mit Hypophysenadenomoperationen.

BACKGROUND: Pituitary tumors are adenomas of a region of the sella turcica which can produce compression of the anterior visual pathway. PATIENTS AND METHODS: Besides clinical signs of visual improvement such as enhanced visual acuity and visual field, the use of electroophthalmological methods can help monitor patients after neurosurgery. RESULTS: The N75 P100 amplitude of the p-VEP has proven to be the most sensitive marker for measuring-improvements after surgery. This amplitude significantly increased, whereas latency time, visual acuity and visual field showed no statistically significant changes after surgery. CONCLUSIONS: The p-VEP is an important tool in treating patients with compression of the visual pathway.

Rational design of an improved induction scheme for recombinant Escherichia coli.

In Escherichia coli, strong overexpression of a recombinant protein has been shown to be deleterious due to a heavy metabolic burden on the host cell, which may completely cease cell growth before maximum product accumulation has occurred. Aiming at a reduction of very high product formation rates, we engineered E. coli strains by mutating the Leloir pathway for galactose metabolization, so that galactose can be utilized to induce lac derived promoters. The induction with galactose was effective in every strain and expression construct tested, and it reduced the metabolic burden on a highly overproducing clone so that cell growth and product accumulation could be maintained for several generations.

[Is cranial corneoscleral 6 mm "no-stitch" tunnel incision contraindicated in against-the-rule astigmatism?]. / Ist eine kraniale korneosklerale 6 mm "No-stitch"-Tunnelinzision bei einem Astigmatismus gegen die Regel kontraindiziert?

BACKGROUND: There is a risk to increase a preexisting astigmatism against the rule by performing the incision at the 12 o'clock position. PATIENTS AND METHODS: In the prospective study 15 patients with a preoperative astigmatism over 0.25 D axis 90 degrees +/- 15 degrees the first cataract extraction was performed by a cranial, the operation of the other eye by a temporal 6-mm no-stitch tunnel incision. RESULTS: The mean corneal astigmatism against the rule preoperatively was 1.02 +/- 0.5 D with cranial incision and 1.27 +/- 0.8 D with temporal incision. Postoperatively the corneal astigmatism was 2.18 +/- 0.9 D with cranial incision and 0.71 +/- 0.7 D with temporal incision. In comparison to the pre-existing astigmatism, there was an increase of the astigmatism with cranial incision more than double, with temporal incision the astigmatism was significant lowered. Postoperatively the astigmatism with temporal wound location was significant lower than with cranial incision. CONCLUSION: Under the aspect of a 1.5 D higher average astigmatism postoperatively, we conclude that a cranial 6-mm no-stitch tunnel incision in preoperative astigmatism against the rule is contraindicated.

Optimization of recombinant gene expression in Escherichia coli.

The major targets for improvement of recombinant expression efficiency in Escherichia coli are gene dosage, transcription and, to some extent, translation. In order to evaluate the relative importance of these factors, the kinetics of specific mRNA compared to product formation was studied for different widely used expression systems, producing recombinant human superoxide dismutase. For a system employing phage T7 RNA polymerase, where a high level of recombinant protein expression puts a high metabolic burden on the cells, it was shown that transcription is not the limiting factor. To improve the translation rate of a common vector based on the tac promoter, the Shine-Dalgarno (SD) sequence was mutated towards stronger homology to the anti-SD sequence of the E. coli 16S rRNA. A 12.2-fold increase in protein yield was accompanied by a 4.3-fold increase in specific mRNA, indicating that transcription of the recombinant gene is coupled to translation. As this coupling amplifies the detrimental effect of a low-efficiency ribosomal binding site, much attention should be paid to translation initiation when optimizing a recombinant protein production system. Finally, reasons for the high expression level before induction are discussed, and first results towards reducing it are presented.

[Value of electrophysiology in ophthalmologic monitoring of patients with endocrine orbitopathy]. / Bedeutung der Elektrophysiologie bei der ophthalmologischen Uberwachung von Patienten mit endokriner Orbitopathie.

BACKGROUND: Patients with Graves' ophthalmopathy may suffer from optic nerve involvement. Different ophthalmological examinations reveal the stage of this disease. PATIENTS AND METHODS: In a retrospective study 27 patients with Graves' ophthalmopathy were examined. Best corrected visual acuity, intraocular pressure, visual field, motility and thickness of ocular muscle were determined. Registration of visual evoked cortical potentials (P-VEP) and pattern electroretinograms (P-ERG) were performed regularly. RESULTS: In our study group severe damage of the optic nerve could be avoided by short examination intervals and early treatment. CONCLUSIONS: Regular controls of P-VEP reveal optic nerve involvement at an early stage, at which visual acuity might not be altered. Therefore, this electrophysiological examination should be used in controlling patients suffering from Graves' ophthalmopathy.

[Dapiprazol antagonizes tropicamide- and phenylephrine-induced mydriasis in the elderly]. / Dapiprazol antagonisiert die Tropicamid- und die Phenylephrin-induzierte Mydriasis bei älteren Personen.

Dapiprazole eye drops were applied 1 h after topical tropicamide or phenylephrine instillation in 30 elderly subjects. Pupil diameter normalized much earlier than in controls receiving placebo. Dapiprazole constricted the tropicamide-dilated pupil by 1 mm after 1 h and normalized the pupil diameter of the phenylephrine-dilated pupil within 1 h, whereas the pupils of the patients receiving placebo remained dilated. Intraocular pressure did not change; no major side effects were seen.

Internal Ca2+ ions inactivate and modify ATP-sensitive potassium channels in adult mouse skeletal muscle.

The effects of internal Ca2+ ions on single ATP-sensitive potassium channels (KATP channels) were studied in inside-out membrane patches excised from mouse skeletal muscle. Channel activity was high when patches were excised in a Ca(2+)-free, Na(+)-rich solution and declined irreversibly within seconds in the presence of internal Ca2+ (0.1 and 2 mM). After Ca(2+)-dependent inactivation of the channels, the ATP concentration-response curve was steeper and 50% channel blockage occurred at a lower ATP concentration than before inactivation. ATP (50 microM) in a K(+)-rich solution bathing the intracellular membrane surface reduced the open-probability of KATP channels to 18% before and to 10% after exposure of the patch to internal Ca2+ (0.1 mM). The block of KATP channels by ATP (50 microM) was also enhanced by internal Ca2+ at a concentration of 13 microM. It is concluded that internal Ca2+ ions can both inactivate KATP channels and modify the active channels. KATP channels modified by Ca2+ are blocked more strongly by ATP than unmodified channels.

Isoproterenol enhances a calcium-independent potassium current in mouse anterior pituitary tumor cells.

1. The patch-clamp technique was used to study the action of the beta-adrenergic agonist (-)-isoproterenol in anterior pituitary tumor cells of the mouse. 2. (-)-Isoproterenol induced an inward-rectifying potassium conductance with half-maximal stimulation at a concentration of approximately 67 nM. The isomer (+)-isoproterenol was less effective in stimulating the current. 3. The effect of (-)-isoproterenol was abolished by cholera toxin treatment, indicating the involvement of a Gs protein, whereas pertussis toxin treatment did not exhibit a current reduction. 4. We blocked or stimulated phosphorylation pathways in cells to test the involvement of adenosine 3',5'-cyclic monophosphate (cAMP). It was concluded that the current stimulation probably was not exclusively mediated by cAMP. 5. Activation of calcium-dependent potassium channels by an isoproterenol-induced calcium influx into the cell could be excluded. 6. Therefore it is suggested that the observed activation of a potassium current by isoproterenol could be directly mediated by a Gs protein.

Vanadate as an activator of ATP--sensitive potassium channels in mouse skeletal muscle.

The inside-out mode of the patch-clamp technique was used to study adenosine-5'-triphosphate (ATP)-sensitive K+ channels in mammalian skeletal muscle. Vanadate, applied to the cytoplasmic face of excised patches, was a potent activator of ATP-sensitive K+ channels. Divalent cations (Mg2+, Ca2+) were a prerequisite for the activating process. The maximal effect was achieved using 1 mM vanadate dissolved in Ringer, increasing the open-state probability about ninefold. The active 5 + redox form of vanadate which stimulates ATP-sensitive K+ channels is likely to be decavanadate V10O28(6-). ATP concentration-response curves have Hill coefficients near three internal Na(+)-rich Ringer and between one and two in internal KCl solutions. Half-maximal channel blockage was observed at ATP concentrations of 4 and 8 microM in Ringer and KCl solutions, respectively. Internal vanadate shifted the curves towards higher ATP concentrations without affecting their slopes. Thus 50% channel blockage occurred at 65 microM ATP in internal Ringer containing 0.5 mM vanadate. The results indicate that the affinity and stoichiometry of ATP binding to ATP-sensitive K+ channels are strongly modulated by internal cations and that the ATP sensitivity is weakened by vanadate.

Effects of potassium channel openers on single potassium channels in mouse skeletal muscle.

The patch-clamp technique was used to study the effects of the potassium channel openers cromakalim, pinacidil, RP 49356 and diazoxide on single potassium channels in mouse skeletal muscle. In excised patches in the inside-out configuration, one type of potassium channel, the ATP-sensitive potassium channel, could be activated by internally applied RP 49356 even in the absence of internal ATP. At a concentration of 0.4 and 0.8 mmol/l, RP 49356 increased the open-probability of the channels by a factor of 2.7 and 17.4 respectively. The stimulating effect of cromakalim (0.2-0.8 mmol/l) and pinacidil (0.4 mmol/l) depended on the presence of ATP (0.1 mmol/l) at the cytoplasmic side of the patch membrane. The two drugs were able to restore the open-probability of the channels blocked by internal ATP (0.1 mmol/l) to 50-90% of its value in ATP-free solution. No channel reactivation could be observed at a higher ATP concentration (1 mmol/l). Diazoxide (0.4 mmol/l) had almost no effect. None of these channel openers could stimulate the other prominent type of potassium channel in skeletal muscle, the large-conductance Ca2(+)-activated potassium channel. The results show that cromakalim, pinacidil and RP 49356 are specific openers of ATP-sensitive potassium channels in skeletal muscle. It is suggested that the drugs displace the channel blocker ATP and that RP 49356 in addition recruits inactive channels.

Polymyxin B as a highly effective gating modifier of high-conductance Ca2(+)-activated K+ channels in mouse skeletal muscle.

Large-conductance Ca2(+)-activated K+ channels were studied in excised inside-out membrane patches from adult mouse skeletal muscle. The channels had a conductance of about 250 pS in symmetrical 155 mM KCl solutions. They showed gating characteristics similar to those described for this type of channel in rat and rabbit skeletal muscle. Polymyxin B, a cyclic polypeptide antibiotic, produced a voltage-dependent block, whereas polymyxin E was only slightly effective. The concentration at which half-maximal blockage occurred was very low: 0.5 microgram/ml at a voltage of +30 mV. The blockage occurred with a Hill coefficient of h = 1.2. At negative membrane potentials, polymyxin B caused the appearance of a substate with a conductance of about 10% of the fully open state. The mode of blockage is discussed and compared to the effect of polymyxin B on glucose uptake into the muscle cell.

ATP-sensitive potassium channels in adult mouse skeletal muscle: characterization of the ATP-binding site.

Single K+-selective channels were studied in excised inside-out membrane patches from dissociated mouse toe muscle fibers. Channels of 74 pS conductance in symmetrical 160 mM KCl solutions were blocked reversibly by 10 microM internal ATP and thus identified as ATP-sensitive K+ channels. The channels were also blocked reversibly by mM concentrations of internal adenosine, adenine and thymine, but not by cytosine and uracil. The efficacy of the reversible channel blockers was higher when they were present in internal NaCl instead of KCl solutions. An irreversible inhibition of ATP-sensitive K+ channels was observed after application of several sulphydryl-modifying substances in the internal solution: 0.5 mM chloramine-T, 50 mM hydrogen peroxide or 2 mM N-ethylmaleimide (NEM). Large-conductance Ca-activated K+ channels were not affected by these reagents. The presence of 1 mM internal ATP prevents the irreversible inhibition of ATP-sensitive K+ channels by NEM. The results suggest that internal Na+ ions increase the affinity of the ATP-sensitive K+ channel to ATP and to other reversible channel blockers and that a functionally important SH-group is located at or near the ATP-binding site.

Low-conductance states of K+ channels in adult mouse skeletal muscle.

Single-channel currents were recorded from Ca2+-activated or ATP-sensitive K+ channels in inside-out membrane patches excised from isolated mouse toe muscles. In addition to the closed and fully open configurations, both types of channels may exhibit several intermediate low-conductance states which are clustered near multiples of elementary conductance units. The units are 1/8 or 1/6 of the channel conductance for Ca2+-activated channels and 1/4 or 1/3 for ATP-sensitive channels. Normally, low-conductance states are rare, but they occur more frequently directly after patch excision. An increased probability of low-conductance states of ATP-sensitive K+ channels was also observed in the presence and during washout of the internal channel blocker adenine. The results suggest that Ca2+-activated and ATP-sensitive K+ channels are composed of several membrane pores with strong positive cooperativity among the elementary conductance units.

Niacin requirement for growth of axenic Entamoeba histolytica.

Niacin (nicotinic acid) was found to be essential for the cultivation of axenic Entamoeba histolytica. This vitamin requirement was also satisfied by nicotinamide. Panmede liver digest, a source of vitamins in the axenic medium was replaced with a dialyzed hot water extract of homogenized whole liver, supplemented with various growth factors. Cultured medium made with the liver extract and not supplemented with niacin failed to support continued multiplication of E. histolytica, but did support serial subculture when niacin was added. The concentration of added niacin required to achieve maximum growth was about 1 microgram per ml of medium. This is the first demonstration of a niacin requirement by the organism.