Compound heterozygous variants in NBAS as a cause of atypical osteogenesis imperfecta.

BACKGROUND: Osteogenesis imperfecta (OI), the commonest inherited bone fragility disorder, affects 1 in 15,000 live births resulting in frequent fractures and reduced mobility, with significant impact on quality of life. Early diagnosis is important, as therapeutic advances can lead to improved clinical outcome and patient benefit. REPORT: Whole exome sequencing in patients with OI identified, in two patients with a multi-system phenotype, compound heterozygous variants in NBAS (neuroblastoma amplified sequence). Patient 1: NBAS c.5741G>A p.(Arg1914His); c.3010C>T p.(Arg1004*) in a 10-year old boy with significant short stature, bone fragility requiring treatment with bisphosphonates, developmental delay and immunodeficiency. Patient 2: NBAS c.5741G>A p.(Arg1914His); c.2032C>T p.(Gln678*) in a 5-year old boy with similar presenting features, bone fragility, mild developmental delay, abnormal liver function tests and immunodeficiency. DISCUSSION: Homozygous missense NBAS variants cause SOPH syndrome (short stature; optic atrophy; Pelger-Huet anomaly), the same missense variant was found in our patients on one allele and a nonsense variant in the other allele. Recent literature suggests a multi-system phenotype. In this study, patient fibroblasts have shown reduced collagen expression, compared to control cells and RNAseq studies, in bone cells show that NBAS is expressed in osteoblasts and osteocytes of rodents and primates. These findings provide proof-of-concept that NBAS mutations have mechanistic effects in bone, and that NBAS variants are a novel cause of bone fragility, which is distinguishable from 'Classical' OI. CONCLUSIONS: Here we report on variants in NBAS, as a cause of bone fragility in humans, and expand the phenotypic spectrum associated with NBAS. We explore the mechanism underlying NBAS and the striking skeletal phenotype in our patients.

Direct stimulation of bovine ovarian progesterone secretion by low concentrations of alpha-interferon.

The ruminant conceptus secretes proteins during early pregnancy which maintain the corpus luteum. These trophoblast proteins are related to the alpha II-interferons and prevent luteolysis indirectly by disrupting the secretion of endometrial prostaglandin. Although trophoblast interferons appear to be largely confined to the uterine lumen, it remains possible that they also act peripherally. This report describes in vitro studies which suggest that interferon may influence hormone secretion by the ovary directly. The study employed i) a well defined serum-free culture model in which bovine granulosa cells secrete the luteal hormones progesterone and oxytocin, and ii) serum-free and serum-supplemented cultures of cells from early CL. Dose-response experiments were performed using bovine recombinant alpha-interferon (brIFN). Progesterone and oxytocin secretions were measured over 4-5 days of culture and DNA content was also determined. Low concentrations of brIFN (10(-15) mol/l to 10(-11) mol/l) stimulated progesterone secretion by granulosa cells by up to three fold, without significantly affecting oxytocin concentrations or culture DNA content. Concentrations of 10(-10) mol/l to 10(-1) mol/l suppressed progesterone secretion in a log dose-related manner (r = 0.97) with evidence of toxicity (lower oxytocin concentrations and significantly reduced DNA compared with controls). Progesterone secretion by luteal cells in serum-free culture was stimulated in the presence of 10(-15) mol/l brIFN, whilst high concentrations again caused inhibition. The data show that ovarian cells can respond directly to low concentrations of interferon-like proteins.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of oxytocin and oestradiol on the action of conceptus secretory protein in progesterone-treated ovariectomized ewes.

In intact cyclic ewes intrauterine infusion of conceptus secretory proteins results in the suppression of both endometrial oxytocin receptor concentrations and oxytocin-induced prostaglandin F2 alpha release. However, similar infusion in progesterone-treated ovariectomized ewes, while suppressing endometrial oxytocin receptors, does not fully inhibit oxytocin-induced prostaglandin F2 alpha release. To examine whether this anomaly resulted from an inadequate simulation of the luteal phase in the ovariectomized ewe treated with progesterone alone, the effects of additional treatment with two other ovarian hormones, oestradiol-17 beta and oxytocin, was investigated. Rather than permitting conceptus secretory protein to successfully inhibit oxytocin-induced prostaglandin F2 alpha release, treatment with oestradiol-17 beta in addition to progesterone actually resulted in an advancement in the timing of release. However, treatment with oxytocin, alone or in combination with oestradiol, permitted the full inhibition of oxytocin-induced prostaglandin F2 alpha release. To confirm that this effect did not result from the action of oxytocin alone, independently of the action of conceptus secretory protein, a second experiment was undertaken using a similar protocol but without the infusion of conceptus secretory protein. In this situation, oxytocin-induced prostaglandin F2 alpha release was only partially inhibited suggesting that both luteal oxytocin and conceptus secretory proteins are necessary to facilitate the full inhibition of luteolysis during early pregnancy in the ewe.

Regulation of oxytocin, oestradiol and progesterone receptor concentrations in different uterine regions by oestradiol, progesterone and oxytocin in ovariectomized ewes.

The regulation of oxytocin, oestradiol and progesterone receptors in different uterine cell types was studied in ovariectomized ewes. Animals were pretreated with a progestogen sponge for 10 days followed by 2 days of high-dose oestradiol to simulate oestrus. They then received either low-dose oestradiol (Group E), low-dose oestradiol plus progesterone (Group P) or low-dose oestradiol, progesterone and oxytocin (via osmotic minipump; Group OT). Animals (three to six per time-point) were killed following ovariectomy (Group OVX), at oestrus (Group O) or following 8, 10, 12 or 14 days of E, P or OT treatment. In a final group, oxytocin was withdrawn on day 12 and ewes were killed on day 14 (Group OTW). Oxytocin receptor concentrations and localization in the endometrium and myometrium were measured by radioreceptor assay, in situ hybridization and autoradiography with the iodinated oxytocin receptor antagonist d(CH2)5[Tyr(Me)2,Thr4,Tyr-NH2(9)]-vasotocin. Oestradiol and progesterone receptors were localized by immunocytochemistry. Oxytocin receptors were present in the luminal epithelium and superficial glands of ovariectomized ewes. In Group O, endometrial oxytocin receptor concentrations were high (1346 +/- 379 fmol [3H]oxytocin bound mg protein-1) and receptors were also located in the deep glands and caruncular stroma in a pattern resembling that found at natural oestrus. Continuing low-dose oestradiol was unable to sustain high endometrial oxytocin receptor concentrations with values decreasing significantly to 140 +/- 20 fmol mg protein-1 (P < 0.01), localized to the luminal epithelium and caruncular stroma but not the glands. Progesterone treatment initially abolished all oxytocin receptors with none present on days 8 or 10. They reappeared in the luminal epithelium only between days 12 and 14 to give an overall concentration of 306 +/- 50 fmol mg protein-1. Oxytocin treatment caused a small increase in oxytocin receptor concentration in the luminal epithelium on days 8 and 10 (20 +/- 4 in Group P and 107 +/- 35 fmol mg protein-1 in Group OT, P < 0.01) but the rise on day 14 was not affected (267 +/- 82 in Group OT and 411 +/- 120 fmol mg protein-1 in Group OTW). In contrast, oestradiol treatment was able to sustain myometrial oxytocin receptors (635 +/- 277 fmol mg protein-1 in Group O and 255 +/- 36 in Group E) and there was no increase over time in Groups P, OT and OTW with values of 61 +/- 18, 88 +/- 53 and 114 +/- 76 fmol mg protein-1 respectively (combined values for days 8-14). Oestradiol receptor concentrations were high in all uterine regions in Group O. This pattern and concentration was maintained in Group E. In all progesterone-treated ewes, oestradiol receptor concentrations were lower in all regions at all time-points. The only time-related change occurred in the luminal epithelium in which oestradiol receptors were undetectable on day 8 but developed by day 10 of progesterone treatment. Progesterone receptors were present at moderate concentrations in the deep glands, caruncular stroma, deep stroma and myometrium in Group O. Oestradiol increased progesterone receptors in the luminal epithelium, superficial glands, deep stroma and myometrium. Progesterone caused the loss of its own receptor from the luminal epithelium and superficial glands and decreased its receptor concentration in the deep stroma and myometrium at all time-points. There was a time-related loss of progesterone receptors from the deep glands of progesterone-treated ewes between days 8 and 14. These results show differences in the regulation of receptors between uterine regions. In particular loss of the negative inhibition by progesterone on the oxytocin receptor by day 14 occurred only in the luminal epithelium, but is unlikely to be a direct effect of progesterone as no progesterone receptors were present on luminal epithelial cells between days 8 and 14.

Traumatic diaphragmatic hernia.

The records of 36 patients with traumatic diaphragmatic hernia (TDH) were reviewed. In 14, acute hernias were diagnosed, but the diagnosis was made one month to 15 years after injury in 22 patients with chronic hernia. Seven acute TDHs were due to blunt and seven to penetrating trauma. Four chronic TDHs were due to blunt and 18 to penetrating trauma. Chest pain, abdominal pain, or dyspnea occurred in each acute case and in 18 of 22 chronic cases. Plain chest roentgenograms were abnormal in 33 of 36 cases. Pleural effusion or abnormal diaphragmatic contour were common abnormal findings. Supradiaphragmatic bowel, pathognomonic of TDH, was evident in seven acute and eight chronic hernias. Celiotomy was routinely employed in acute hernias, celiotomy or thoracotomy in chronic hernias. There were three deaths, two with associated CNS injury and one with chronic pulmonary disease.

The ultrasonic dissector facilitates laparoscopic cholecystectomy.

The ultrasonic dissector disrupts tissues in proportion to their fluid content by ultrasonically induced cavitational forces. Since sturdy tissues are spared, the instrument tends to follow tissue planes and to dissect fat and other soft tissues selectively. We performed a prospective, randomized, controlled trial in 73 patients comparing the safety and efficacy of a prototype ultrasonic dissector with that of electrosurgery and laser during laparoscopic cholecystectomy. Randomization was as follows: ultrasonic dissector, 37 patients; electrosurgery, 21 patients; and laser, 15 patients. The results were not different with respect to patient characteristics, amount of blood loss, technical difficulties, length of hospital stay, or return to work. Subjectively, the ultrasonic dissector was thought to be of special value in isolating the hilar structures, particularly when they were edematous or embedded in fat. The ultrasonic dissector disintegrated the fat, which was rapidly cleared up the suction channel, allowing the cystic duct and artery to be bared with less risk of injury. We concluded that the ultrasonic dissector has unique attributes that contribute to the ease and safety of laparoscopic cholecystectomy.

Laparoscopic or open inguinal herniorrhaphy? A randomized prospective trial.

OBJECTIVE: To determine whether transabdominal preperitoneal laparoscopic hernia repair can equal or surpass an established open method at an acceptable cost. DESIGN: A randomized, prospective comparison with a follow-up of 7 to 18 months (median, 10 months; planned, 5 years). SETTING: Health maintenance organization hospital. PATIENTS: One hundred patients between 20 and 70 years of age were randomized. No patient withdrew from the study after randomization. INTERVENTIONS: Transabdominal preperitoneal laparoscopic and open tension-free repairs using a polypropylene mesh. MAIN OUTCOME MEASURES: Operative and discharge times, costs, recovery, and morbidity. "Return to work" was supplemented by a performance assessment using a panel of exercises. RESULTS: Operative and hospitalization times were not significantly different between the two types of repair. Patients with laparoscopic unilateral repairs returned to work faster (9 vs 17 days). At 1 week postoperatively, performance of straight-leg raises correlated well with time to return to work for patients with strenuous jobs. The laparoscopic repair was more expensive than the open approach ($3093 vs $2494). CONCLUSIONS: Laparoscopic transabdominal preperitoneal hernia repair can be accomplished with operative and hospitalization times and a short-term recurrence rate similar to those of an established open technique. Perioperative exercise testing may be an important adjunct to return to work in the comparison of methods.

Hormonal regulation of oxytocin-induced prostaglandin F(2alpha) secretion by the bovine and ovine uterus in vivo.

In long-term ovariectomized ewes and cows, endometrial oxytocin receptors rest at relatively high levels but oxytocin is unable to induce prostaglandin F(2alpha) release. A series of studies were carried out to investigate the roles of physiological levels of progesterone and estradiol in "activating" these receptors in terms of permitting oxytocin-induced prostaglandin F(2alpha) release. In long-term ovariectomized cows, treatment with progesterone, but not estradiol, resulted in the induction of responsiveness to oxytocin. This responsiveness appeared within 2 d of progesterone treatment, reached a maximum by 6 d and was maintained to Day 18. In ovariectomized ewes, while estradiol treatment did induce temporary responsiveness to oxytocin after 3 d of treatment, treatment with progesterone was required to induce sustained responsiveness that appeared by Day 9 of treatment and was maintained to Day 12. Measurement of endometrial receptors for oxytocin revealed a significant decline in oxytocin receptors by Day 6 of progesterone treatment when responsiveness to oxytocin was maximal, demonstrating that receptor concentrations were not a limiting factor. The most likely mechanism by which progesterone treatment induces responsiveness to oxytocin may be through the up regulation of post receptor signaling pathways and/or enzymes involved in prostaglandin synthesis.

The effects of adrenaline and atropine on oxytocin-induced estrus in the goat.

Oxytocin-induced luteolysis in goats was associated with significant increases in peripheral plasma concentrations of 13,14-dihydro-15-keto-prostaglandin (PG) F(2alpha) (PGFM). This effect was not inhibited by concomitant administration of adrenaline (1 mg), although increases of PGFM were both delayed and diminished. Administration of atropine, a muscarinic cholinergic antagonist, inhibited the effect of oxytocin in three out of five goats. In these animals, increases in PGFM were inhibited.

Effect of adrenalin and propranolol on progesterone and oxytocin secretion in vivo during the caprine estrous cycle.

The effects of jugular infusions of adrenalin and the beta-adrenergic receptor antagonist propranolol on plasma concentrations of progesterone and oxytocin were examined at 2 different stages of the caprine estrous cycle. Adrenalin (25 micrograms.kg-1h-1) significantly (P < 0.05) increased oxytocin secretion on Day 3 and Day 10 of the cycle (estrus = Day 0); progesterone concentrations were significantly (P < 0.05) elevated on Day 10 alone. Propranolol had no effect on progesterone secretion yet significantly (P < 0.05) reduced oxytocin concentrations on Day 3. These results suggest that there may be neuroendocrine involvement in the regulation of luteal oxytocin secretion in the goat.

Effects of arterial infusions of adrenalin and acetylcholine on luteal secretion of progesterone and oxytocin in goats.

The effects of close intra-arterial infusion of acetylcholine and adrenalin on ovarian secretion of progesterone and oxytocin were examined on Day 10 of the estrous cycle in goats (estrus = Day 0). Acetylcholine (15 micrograms/min) was without effect, but adrenalin (10 micrograms/min) significantly (P < 0.001) raised both progesterone and oxytocin concentrations in ovarian vein plasma. These results show that luteal hormone secretion is enhanced in the goat by beta-adrenergic stimulation and suggest that, as in the sheep and cow, there may be neuroendocrine involvement in the regulation of caprine luteal function.

Insensitivity of dispersed caprine luteal cells to beta-adrenergic agonists and other putative transmitter substances.

Adrenaline (10(-6)-10(-4)M), serotonin (10(-6)-10(-4)M), and several other potential steroidogenic agonists failed to enhance either basal or LH-stimulated progesterone production from dispersed caprine luteal cells from Day 10 of the estrous cycle. The caprine corpus luteum (CL) would appear to be more refractory to exogenous stimulation than either the ovine or bovine CL.

Pulsatile release of oxytocin after suppression of prostaglandin synthesis in hysterectomised ewes.

On days 13 and 14 after oestrus (day 0) oxytocin and 13,14-dihydro-15-keto-prostaglandin (PG)F2 alpha (PGFM) concentrations were measured in jugular plasma of hysterectomised sheep with or without systemic treatment with the PG cyclo-oxygenase inhibitor indomethacin (4 mg kg-1 three times a day on days 12, 13 and 14 subcutaneously. Pulsatile increases of oxytocin were observed in both untreated and treated sheep with mean (+/- SD) peak heights of 18.4 +/- 9.6 pg ml-1 (n = 11) and 23.5 +/- 9.4 pg ml-1 (n = 8), respectively; these means were not significantly different. Plasma concentrations of PGFM remained consistently low in both groups (under 100 pg ml-1) with no significant peaks observed. The data suggest that PGF2 alpha may not be the only stimulus for the release of luteal oxytocin, or that there may be a contribution by the posterior pituitary to oxytocin secretion during the luteal phase of the ovine oestrous cycle.

Endoscopic surgery teleconferencing.

Modern telecommunication allowing the transmission of digitalized voice and images through telephone lines has expanded to medicine. The Integrated Digital Network System (ISDN) can be used to transmit 128 kbit/sec (ISDN-2) to 2 Mbit/sec (ISDN-30). Together with smart compression techniques a teleconferencing system can be implemented to transmit images of an endoscopic operation. In this paper 2 such experiments are described. In the first experiment a laparoscopic cholecystectomy performed in the Netherlands is followed at the same time by surgeons in Hawaii through ISDN-2 lines. In the second experiment a laparoscopic hernia repair was followed in Orlando during a congress using ISDN-6 lines. The images sent over the ISDN-2 lines are rather blurry with rapid movement. The images over the ISDN-6 lines were more acceptable. Data compression and decompression results in a time delay of approximately 1 second, which seems not disturbing during the connection. The cost of communication is comparable to telephone communication, which makes this form of telecommunication feasible in surgical practice.

Prevalence of faecal excretion of verocytotoxigenic Escherichia coli O157 in cattle in England and Wales.

During the decade to 1999, the incidence of human infections with the zoonotic pathogen verocytotoxin-producing Escherichia coli O157 (VTEC O157) increased in England and Wales. This paper describes the results of a survey of 75 farms to determine the prevalence of faecal excretion of VTEC O157 by cattle, its primary reservoir host, in England and Wales. Faecal samples were collected from 4663 cattle between June and December 1999. The prevalence of excretion by individual cattle was 4.2 per cent (95 per cent confidence interval [CI] 2.0 to 6.4) and 10.3 per cent (95 per cent CI 5.8 to 14.8) among animals in infected herds. The within-herd prevalence on positive farms ranged from 1.1 to 51.4 per cent. At least one positive animal was identified on 29 (38.7 per cent; 95 per cent CI 28.1 to 50.4) of the farms, including dairy, suckler and fattening herds. The prevalence of excretion was least in the calves under two months of age, peaked in the calves aged between two and six months and declined thereafter. The phage types identified most widely were 4, 34 and 2, which were each found on six of the 29 positive farms.

Teleconferencing bridges two oceans and shrinks the surgical world.

Endoscopic surgery has led to changes in surgical practice which may rival the introduction of anesthesia and antibiotics in significance. As a result, an exciting synergy has rapidly emerged between technology and clinical practice. However, questions of training, credentialing, and patient safety have been raised as traditional procedures have been adapted to the minimally invasive approach and new ones are described. Many surgeons have been reluctant to venture beyond laparoscopic cholecystectomy. Halting first efforts at advanced procedures may prolong operative times, increase risk, and raise costs. Older methods of surgical education are not adequate to meet the current need (Fig. 1).

The marginal seal of Class II restorations: flowable composite resin compared to injectable glass ionomer.

The objective of this study was to investigate the effect of a flowable composite resin (Tetric Flow) versus an injectable glass ionomer (Fuji II LC) on microleakage at the cavosurface margin of the proximal box of Class II restorations in permanent teeth in-vitro. Thirty caries and restoration-free human bicuspids were prepared with mesial and distal slot preparations and were filled either with a bonding agent (Optibond) plus a flowable composite resin (Tetric Flow), Group I; bonding agent (Optibond) plus a flowable glass ionomer (Fuji II LC), Group II; or a flowable glass ionomer (Fuji II LC) with no bonding agent, Group III. All specimens were then immersed in a 2% solution of basic fuschin dye for 24 hours to allow for dye penetration into possible existing gaps. These teeth were then carefully sectioned mesially/distally into two pieces using an Isomet saw. The teeth were then studied under a binocular microscope to measure depths of dye penetrations as an indication of marginal microleakage at the gingival cavosurface margin and scored as follows: 0 = no dye penetration, 1 = dye penetration into enamel only, 2 = dye penetration into enamel and dentin, 3 = dye penetration into the pulp. The specimens were also evaluated using a SEM. The results showed that there were statistically significant differences between Groups I (Tetric Flow) and (Fuji II LC plus bonding agent), II in favor of Group I; between Groups I and (Fuji II LC with no bonding agent), III in favor of Group I; as well as Groups II and III in favor of Group II (Fuji II LC plus bonding agent). Group I (bonding agent plus flowable composite resin) showed significantly less microleakage. Group II (bonding agent plus flowable glass ionomer) demonstrated a bond that existed between the bonding agent and the glass ionomer but microleakage within microgaps of the glass ionomer itself Group III (flowable glass ionomer plus no bonding agent) demonstrated significant microleakage between the glass ionomer and tooth structure, microgaps within the glass ionomer, and lack of retention of the restoration. It appears that the use of a flowable composite resin (Tetric Flow) plus a bonding agent (Optibond) in the proximal box of a Class II restoration in permanent teeth will significantly reduce the microleakage at the cavosurface margin when compared with an injectable glass ionomer (Fuji II LC) with or without a bonding agent (Optibond).

Laparoscopic staging of malignant disease.

The earliest applications of laparoscopy were for diagnostic procedures. The goal was to make a diagnosis while sparing the patient a major, and often futile, operation. However, the view was limited and it was not possible to palpate organs or masses. The recent development of advanced laparoscopic techniques and ultrasound have improved our view and restored our "sense of touch". These innovations bring the goal of minimally invasive diagnosis and staging closer to reality. This paper reviews the current literature on the laparoscopic staging of cancer with an emphasis on patient selection, diagnostic accuracy, and the reduction in morbidity which can be achieved.

Advanced laparoscopy: "the next generation". The adrenal, kidney, spleen, pancreas, and liver.

Following the rapid acceptance of laparoscopic cholecystectomy, a few adventurous surgeons "dared to boldly go where no one had gone before." They sought to achieve the same reduction in morbidity while accomplishing the same surgical goals. This paper will briefly review the current status of minimal access surgery for the solid organs of the abdomen. It will focus on the indications, risks, limitations, and on the balance between the trauma of access and the trauma of the procedure itself. As new techniques and equipment emerge and experience and data are accumulated, this balance may shift. Some of these procedures are in their infancy while others are rapidly becoming the new "Gold Standard".

Case study of copper poisoning in a British dairy herd.

Following the initial diagnosis of chronic copper poisoning (CCP), the copper (Cu) status of a British dairy herd was investigated. Eight fatal cases of CCP were identified over a 17-month period, from December 1999 to May 2001, involving seven Jersey cows and one Holstein-Friesian; seven cows were dry when CCP occurred. Case diagnostic criteria were necrotising hepatopathy associated with abnormally high liver and kidney Cu concentrations. Analysis of the ration for the high-yielding Jersey cow group revealed about 50 mg Cu/kg dry matter intake (DMI). Risk factors predisposing to fatal CCP were Jersey breed, previous high yield, first two weeks of the dry period and moderately high dietary Cu (greater than 40 mg Cu/kg DMI).