Gene replacement therapy after neuropathy onset provides therapeutic benefit in a model of CMT1X.

X-linked Charcot-Marie-Tooth disease (CMT1X), one of the commonest forms of inherited demyelinating neuropathy, results from GJB1 gene mutations causing loss of function of the gap junction protein connexin32 (Cx32). The aim of this study was to examine whether delayed gene replacement therapy after the onset of peripheral neuropathy can provide a therapeutic benefit in the Gjb1-null/Cx32 knockout model of CMT1X. After delivery of the LV-Mpz.GJB1 lentiviral vector by a single lumbar intrathecal injection into 6-month-old Gjb1-null mice, we confirmed expression of Cx32 in lumbar roots and sciatic nerves correctly localized at the paranodal myelin areas. Gjb1-null mice treated with LV-Mpz.GJB1 compared with LV-Mpz.Egfp (mock) vector at the age of 6 months showed improved motor performance at 8 and 10 months. Furthermore, treated mice showed increased sciatic nerve conduction velocities, improvement of myelination and reduced inflammation in lumbar roots and peripheral nerves at 10 months of age, along with enhanced quadriceps muscle innervation. Plasma neurofilament light (NEFL) levels, a clinically relevant biomarker, were also ameliorated in fully treated mice. Intrathecal gene delivery after the onset of peripheral neuropathy offers a significant therapeutic benefit in this disease model, providing a proof of principle for treating patients with CMT1X at different ages.

Intrathecal gene therapy in mouse models expressing CMT1X mutations.

Gap junction beta-1 (GJB1) gene mutations affecting the gap junction protein connexin32 (Cx32) cause the X-linked Charcot-Marie-Tooth disease (CMT1X), a common inherited neuropathy. Targeted expression of virally delivered Cx32 in Schwann cells following intrathecal injection of lentiviral vectors in the Cx32 knockout (KO) mouse model of the disease has led to morphological and functional improvement. To examine whether this approach could be effective in CMT1X patients expressing different Cx32 mutants, we treated transgenic Cx32 KO mice expressing the T55I, R75W or N175D CMT1X mutations. All three mutants were localized in the perinuclear compartment of myelinating Schwann cells consistent with retention in the ER (T55I) or Golgi (R75W, N175D) and loss of physiological expression in the non-compact myelin. Following intrathecal delivery of the GJB1 gene we detected the virally delivered wild-type (WT) Cx32 in non-compact myelin of T55I KO mice, but only rarely in N175D KO or R75W KO mice, suggesting dominant-negative effects of the R75W and N175D mutants but not of the T55I mutant on co-expressed WT Cx32. GJB1 treated T55I KO mice showed improved motor performance, lower ratios of abnormally myelinated fibers and reduction of inflammatory cells in spinal roots and peripheral nerves compared with mock-treated littermates. Either partial (N175D KO) or no (R75W KO) improvement was observed in the other two mutant lines. Thus, certain CMT1X mutants may interfere with gene addition therapy for CMT1X. Whereas gene addition can be used for non-interfering CMT1X mutations, further studies will be needed to develop treatments for patients harboring interfering mutations.

Simultaneous occurrence of a medullary and a papillary thyroid carcinoma in the same patient.

We present the case of a patient in whom we diagnosed two different thyroid carcinomas (one on each lobe) of distinct histologic type: one derived from the follicular cells (papillary) and one from the C cells (medullary). They were both diagnosed preoperatively by fine needle aspiration (FNA), and the diagnosis was confirmed with histologic examination. "Inappropriate" staining with neuroendocrine markers was observed in the papillary tumor. Analysis of tumor tissue for the RET oncogene mutations, commonly found in the MEN2 syndromes, was negative. This case supports the view of a common origin for these two tumor types.

Cytogenetic findings in uterine epithelioid leiomyomas.

Epithelioid leiomyomas of the uterus, unlike ordinary leiomyomas, show substantial epithelial differentiation. No chromosome abnormalities have been reported in uterine epithelioid leiomyomas before. We analyzed short-term cultures from five such tumors and detected abnormal karyotypes in four. A del(7) (q21.2q31.2) was found in two tumors, in one as the only change and in the other as a secondary aberration acquired during clonal evolution. Rearrangement of chromosomal band 12q15, another of the cytogenetic hallmarks of ordinary uterine leiomyomas, was seen in the form of a t(10;12) in one tumor. Band 17q21 was involved in structural aberrations in two cases. The data we present indicate that epithelioid leiomyomas are fundamentally similar cytogenetically, and hence presumably also pathogenetically, to the much more common smooth muscle-differentiated uterine myomas. The only differences hinted at are that epithelioid tumors may be karyotypically more complex and more often have rearrangements of 17q21.

Chromosome analysis of uterine adenomyosis. Detection of the leiomyoma-associated del(7q) in three cases.

Adenomyosis is a uterine disease whose defining characteristic is the presence deep in the myometrium of endometrial glands and stroma. The condition is believed to arise from inordinate downward growth by contiguity from the endometrium rather than from in situ metaplasia or neoplasia. No acquired chromosome abnormalities have been associated with adenomyosis before. We analyzed short-term cultures from three cases and detected in all of them a del(7) (q21.2q31.2), a karyotypic anomaly that has hitherto been found repeatedly only in uterine leiomyomas. The cytogenetic similarity to leiomyoma suggests that the del(7q) was present in the mesenchymal or, more precisely, smooth muscle cells of the adenomyosis lesions. The very fact that clonal chromosome abnormalities were present questions whether the prevailing understanding of adenomyosis pathogenesis is adequate; the cytogenetic data would better fit a model of the disease envisioning the intramyometrial endometrial foci as having arisen through a neoplastic process.

The antidiabetic action of somatostatin-28 as assessed by the artificial endocrine pancreas: greater potency than somatostatin-14.

In order to investigate the action of somatostatin-28 (SS-28) on the metabolic homeostasis of insulin-dependent diabetics, we compared its effects to those of somatostatin-14 (SS-14) in terms of insulin sparing, changes in dextrose demands, glucose fluctuations and behavior of growth hormone and glucagon secretion. Eight insulin-dependent subjects were connected to Artificial Endocrine Pancreas (Biostator) for 84 hours during which they received intravenous infusions of either SS-14, SS-28 or isotonic saline in a randomized order, after a steady state of metabolism had been achieved. Five of the patients received SS-28 100 micrograms/h and SS-14 250 micrograms/h for 10 hours and three of them SS-28, 50 micrograms/h and SS-14 250 micrograms/h for 12 hours. Identical doses of both peptides were administered as bolus infusions prior to the continuous ones. Under SS-28 100 micrograms/h and SS-14 250 micrograms/h patients required 13.5 +/- 2.3 and 14.5 +/- 1.9 U of insulin respectively vs 40 +/- 5.6 U under isotonic saline infusion (mean +/- SEM, P less than 0.005 and P less than 0.01). At the same period the apparatus delivered 15 times more dextrose under SS-28 and 20 times more under SS-14. The magnitude of glucose fluctuations diminished from 64.6 +/- 2.47 mg% without to 41.4 +/- 2 mg% under SS-14 (P less than 0.01) and 46 +/- 3.8 mg% under SS-28 (P less than 0.02). Similar changes were observed in the remaining three patients who received SS-28 in the dose of 50 micrograms/h.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of prolonged (6 months) alpha-glucosidase inhibition on blood glucose control and insulin requirements in patients with insulin-dependent diabetes mellitus.

To examine prolonged alpha-glucosidase inhibition on blood glucose control, Acarbose, a potent alpha-glucosidase inhibitor, was administered for six months to insulin-dependent diabetic patients. Acarbose administration significantly diminished postprandial blood glucose increases by 20-30% and reduced insulin requirements by about 40% in these patients. Symptoms related to its use almost disappeared after the first month of treatment. These results suggest that prolonged alpha-glucosidase inhibition improves glucose tolerance in patients with insulin-dependent diabetes mellitus. Thus, an agent like acarbose might be a useful adjunct to insulin in the treatment of diabetic patients.

Differences between somatostatin-28 and somatostatin-14 with respect to their biological effects in healthy humans and acromegalics.

In order to compare the effects of somatostatin-28 (SS-28) with those of somatostatin-14 (SS-14) in humans, we administered both compounds randomly in 5 healthy persons and 3 patients with active acromegaly. Blood glucose, growth hormone, insulin, glucagon, TSH, FSH, LH and prolactin were estimated after arginine, TRH and LHRH stimulation in the normals and without stimulation in the acromegalics. Both substances were administered in doses of 25, 50, 200 and 250 micrograms. Our results indicate that SS-28 is at least 5 times more potent in man than SS-14 as far as inhibition of growth hormone, insulin, glucagon and prolactin secretion is concerned. On the other hand SS-28 is at least 2 times more potent than SS-14 in the inhibition of TSH, FSH and LH. If this difference in potency is calculated on the basis of equimolarity, the action of SS-28 becomes even much greater. According to these findings, SS-28 appears to be either the main hormone and SS-14 a fragment of it with a lesser degree of biologic activity, or the prohormone with special properties.

Biologic activities of biosynthetic human insulin in healthy volunteers and insulin-dependent diabetic patients monitored by the artificial endocrine pancreas.

This study investigates and compares biosynthetic human insulin (BHI) and purified pork insulin, in healthy volunteers and in insulin-dependent diabetic patients, in terms of biologic action, capacity for controlling diabetic patients, and the requirements of the patients on each insulin. The possible importance of this new insulin in the improved long-term control of diabetic patients led to the experimental design of this protocol.