Ventromedial Hypothalamic Nitric Oxide Production Is Necessary for Hypoglycemia Detection and Counterregulation

OBJECTIVE

The response of ventromedial hypothalamic (VMH) glucose-inhibited neurons to decreased glucose is impaired under conditions where the counterregulatory response (CRR) to hypoglycemia is impaired (e.g., recurrent hypoglycemia). This suggests a role for glucose-inhibited neurons in the CRR. We recently showed that decreased glucose increases nitric oxide (NO) production in cultured VMH glucose-inhibited neurons. These in vitro data led us to hypothesize that NO release from VMH glucose-inhibited neurons is critical for the CRR.

RESEARCH DESIGN AND METHODS

The CRR was evaluated in rats and mice in response to acute insulin-induced hypoglycemia and hypoglycemic clamps after modulation of brain NO signaling. The glucose sensitivity of ventromedial nucleus glucose-inhibited neurons was also assessed.

RESULTS

Hypoglycemia increased hypothalamic constitutive NO synthase (NOS) activity and neuronal NOS (nNOS) but not endothelial NOS (eNOS) phosphorylation in rats. Intracerebroventricular and VMH injection of the nonselective NOS inhibitor N^G-monomethyl-l-arginine (l-NMMA) slowed the recovery to euglycemia after hypoglycemia. VMH l-NMMA injection also increased the glucose infusion rate (GIR) and decreased epinephrine secretion during hyperinsulinemic/hypoglycemic clamp in rats. The GIR required to maintain the hypoglycemic plateau was higher in nNOS knockout than wild-type or eNOS knockout mice. Finally, VMH glucose-inhibited neurons were virtually absent in nNOS knockout mice.

CONCLUSIONS

We conclude that VMH NO production is necessary for glucose sensing in glucose-inhibited neurons and full generation of the CRR to hypoglycemia. These data suggest that potentiating NO signaling may improve the defective CRR resulting from recurrent hypoglycemia in patients using intensive insulin therapy.Intensive insulin therapy significantly reduces the onset and progression of hyperglycemia-related complications in patients with type 1 and advanced type 2 diabetes. However, intensive insulin therapy also causes a clinically adverse effect: hypoglycemia (1). Powerful neuroendocrine and autonomic counterregulatory mechanisms protect the brain from hypoglycemia (2,3). These protective mechanisms, known as the counterregulatory response (CRR) to hypoglycemia, involve the release of hormones (e.g., glucagon, epinephrine) that restore euglycemia by stimulating hepatic glucose production and inhibiting peripheral glucose uptake (3). Although the physiology of the CRR is well understood, the underlying cellular mechanisms by which the brain senses hypoglycemia and initiates the CRR remain elusive.During hypoglycemia, central and peripheral glucose sensors detect declining glucose levels (4). In the brain, the ventromedial hypothalamus, which includes the arcuate nucleus and the ventromedial nucleus (VMN), is important in the initiation of the CRR (5–7). This region contains specialized glucose-sensing neurons (GSNs). Ventromedial hypothalamic (VMH) GSN electrical activity is regulated by physiologically relevant changes in extracellular glucose levels (8–11). Glucose-excited neurons decrease, whereas glucose-inhibited neurons increase, their input resistance, membrane potential, and action potential frequency when extracellular glucose is reduced (10). Many studies suggest that VMH glucose-inhibited neurons play a critical role in the control of the CRR (4). For example, the response of VMH glucose-inhibited neurons to decreased glucose is impaired under conditions where the CRR is impaired (e.g., recurrent hypoglycemia) (12,13).Nitric oxide (NO) is a gaseous messenger produced by NO synthase (NOS). Two classes of NOS have been identified in the brain: the inducible NOS (iNOS) and the constitutive NOS, which includes the neuronal NOS (nNOS) and endothelial NOS (eNOS) isoforms (14). Hypothalamic NO is involved in the regulation of food intake and glucose homeostasis (15–18). In support of this, we have recently shown that VMH glucose-inhibited neurons produce NO via nNOS in response to decreased extracellular glucose levels (19,20). Therefore, in this study, we test the hypothesis that NO production by VMH glucose-inhibited neurons is necessary for the CRR to hypoglycemia. We tested this hypothesis using a combination of in vivo and in vitro techniques in wild-type rats and mice as well as in transgenic nNOS and eNOS knockout mice.     

Efficacy of the InVance male sling in men with stress urinary incontinence

OBJECTIVES: To evaluate the efficacy and safety of the InVance bulbourethral sling in male stress urinary incontinence. MATERIALS AND METHODS: Between June 2003 and April 2005, the InVance bulbourethral sling was implanted into 50 patients with urinary incontinence after prostate surgery in 49 cases and pelvic trauma in 1 case. The patients were monitored and evaluated in a prospective manner (continence, tolerance, and satisfaction). The treatment was considered to be successful if the patient stopped wearing any kind of continence pad (patient cured) or only one pad per day (patient improved), with no de novo urinary disorders and without significant postvoid residual urine. Patient satisfaction with the procedure was assessed. RESULTS: After a median follow-up of 6 mo, 50% of patients were dry, 26% had improved, and 24% suffered treatment failure. The success rates for the patients with severe incontinence and those who had undergone radiation therapy were 50% and 25%, respectively. All patients who were dry or had improved were satisfied and presented no obstructive or irritative de novo urinary disorders. The overall success rate for the 51 procedures conducted was 74.5%. Six cases of transitory acute urine retention and six cases of persistent perineal pain were reported. Explantation was necessary because of suppuration of the sling in three patients and of a de novo irritative urinary disorder in one patient. No cases of pubic osteitis or urethral erosion were reported. CONCLUSIONS: The InVance bulbourethral sling procedure makes it possible to treat stress urinary incontinence after prostate surgery with satisfactory and lasting short-term results. Severe incontinence and a past history of pelvic radiation therapy seem to be factors contributing to the failure of this procedure.     

Effects of Intermittent Femoral Nerve Injections of Bupivacaine, Levobupivacaine, and Ropivacaine on Mitochondrial Energy Metabolism and Intracellular Calcium Homeostasis in Rat Psoas Muscle

Background: Long-acting local anesthetics cause muscle damage. Moreover, long-acting local anesthetics act as uncoupler of oxidative phosphorylation in isolated mitochondria and enhance sarcoplasmic reticulum Ca2+ release. The aim of the study was to evaluate effects of perineural injections of local anesthetics on mitochondrial energetic metabolism and intracellular calcium homeostasis in vivo.

Methods: Femoral nerve block catheters were inserted in adult male Wistar rats. Rats were randomized and received seven injections (1 ml/kg) of bupivacaine, levobupivacaine, ropivacaine, or isotonic saline at 8-h intervals. Rats were killed 8 h after the last injection. Psoas muscle was quickly dissected from next to the femoral nerve. Local anesthetic concentrations in muscle were determined. Oxidative capacity was measured in saponin-skinned fibers. Oxygen consumption rates were measured, and mitochondrial adenosine triphosphate synthesis rate was determined. Enzymatic activities of mitochondrial respiratory chain complexes were evaluated. Local calcium release events (calcium sparks) were analyzed as well as sarcoplasmic reticulum calcium content in saponin-skinned fibers.

Results: Eight hours after the last injection, psoas muscle concentration of local anesthetics was less than 0.3 [mu]g/g tissue. Adenosine triphosphate synthesis and adenosine triphosphate-to-oxygen ratio were significantly decreased in the muscle of rats treated with local anesthetics. A global decrease (around 50%) in all of the enzyme activities of the respiratory chain was observed. Levobupivacaine increased the amplitude and frequency of the calcium sparks, whereas lower sarcoplasmic reticulum calcium content was shown.     

XM-ONE detection of endothelium cell antibodies identifies a subgroup of HLA-antibody negative patients undergoing acute rejection

Background

Kidney transplant recipients exhibiting antibodies (Ab) against either HLA or non-HLA antigens undergo frequent episodes of rejection and exhibit decreased long-term graft survival. The novel flow cytometry crossmatch kit XM-ONE, detects Abs to HLA antigens plus those directed to Tie-2-positive precursor endothelial cells (anti-endothelial cell antibodies, AECA). We studied the clinical importance of these lesser known antibodies.

Methods

We retrospectively analyzed 208 sera from 160 recipients of deceased donor grafts for AECA using non-donor peripheral blood endothelial progenitor cells as targets and Luminex methodology for HLA antibodies.

Results

AECA were detected in 64 patients (40%). A significantly higher proportion of patients showing a positive endothelial crossmatch experienced rejection (31 AECA-positive among 43 rejection cases, 72%) compared with those without rejection (33/117, 28.2%). Immunoglobulin M(IgM) predominated (66%) over IgG (14%) and IgG plus IgM (20%). HLA antibodies positively and significantly associated with rejection as expected. Of special interest were the 19 patients who presented with acute rejection episodes along with restricted AECA positivity. The relative-risk for an acute rejection episode with either AECA or HLA—13.87 and 2.43, respectively—was significant. When HLA was already positive, the relative risk for AECA was 1.24, a non-significant increase.

Conclusions

Our data identified AECA-positive patients that showed an increased risk to develop an acute rejection episode early after transplantation. Moreover, restricted AECA-positive patients with acute rejection are an important subgroup which otherwise may be wrongly labeled as non-humoral rejection. Among HLA-negative cases, AECA conferred a significantly greater risk for rejection.     

Absorbing Boundary Conditions for Solving N-Dimensional Stationary Schrödinger Equations with Unbounded Potentials and Nonlinearities

We propose a hierarchy of novel absorbing boundary conditions for the one-dimensional stationary Schrödinger equation with general (linear and nonlinear) potential. The accuracy of the new absorbing boundary conditions is investigated numerically for the computation of energies and ground-states for linear and nonlinear Schrödinger equations. It turns out that these absorbing boundary conditions and their variants lead to a higher accuracy than the usual Dirichlet boundary condition. Finally, we give the extension of these ABCs to N-dimensional stationary Schrödinger equations.     

Pretherapeutic Extraperitoneal Laparoscopic Staging of Bulky or Locally Advanced Cervical Cancer

Background

To assess the safety, feasibility, and impact on survival of extraperitoneal para-aortic lymphadenectomy in the staging of patients with bulky or locally advanced cervical cancer.

Materials and Methods

Between August 2001 and October 2009, 87 consecutive patients (median age 51 years) with bulky or locally advanced cervical cancer underwent extraperitoneal laparoscopic infrarenal aortic and common iliac dissection as a pretherapeutic staging procedure. Data on pathologic findings, details of surgery, postoperative complications, and disease status at follow-up were collected.

Results

The median operating time was 150 min (range 60–255 min). The mean (± standard deviation) para-aortic nodal yield was 15.5 ± 8.1 (range 4–62). In none of the patients, conversion to the transperitoneal approach or laparotomy was necessary. Histological examination revealed metastasis in 13 patients (macroscopic disease 10, microscopic disease 3). After a median follow-up of 33.4 months (range 13.3–65.9 months), 73.6% of patients were free of disease and 1.1% were alive with disease, 19.5% died from cervical cancer, and 3.3% died from other causes. After a follow-up of 3 years, no deaths or recurrences were documented, with an overall survival rate of 74.8% (95% CI 62.8%–83.4%) and disease-free survival of 86% (95% CI 74.7%–92.5%). There were no significant differences in overall survival and disease-free survival between patients with positive and negative para-aortic lymph nodes.

Conclusion

The extraperitoneal laparoscopic para-aortic lymphadenectomy for pretherapeutic surgical staging in cervical cancer is a safe and feasible procedure that should be considered as a tool to identify lymph node positive patients who require extended-field radiation and/or chemotherapy.     

Low pretreatment total testosterone (< 3 ng/mL) predicts extraprostatic disease in prostatectomy specimens from patients with preoperative localized prostate cancer

Study Type – Therapy (case series)
Level of Evidence 4

OBJECTIVE

• ? To investigate the relationship between pretreatment testosterone levels and pathological specimen characteristics, by prospectively examining serum androgen concentrations in a well‐studied cohort of patients who underwent radical prostatectomy (RP) for localized prostate cancer.

PATIENTS AND METHODS

• ? A total of 107 patients with clinically localized prostate cancer had an assay of total testosterone before laparoscopic RP at our institution.
• ? The results were classified into two groups based on the total serum testosterone: group1, <3 ng/mL; group 2, ≥3 ng/mL.
• ? Student’s t‐test was used to compare continuous variables, and Fisher’s exact test or the chi‐squared test was used to compare categorical variables.
• ? Survival curves were established using the Kaplan–Meier method and compared using the log‐rank test. In all tests, P < 0.05 was considered to indicate statistical significance.

RESULTS

• ? All patients had localized prostate cancer based on digital rectal examination (DRE) and preoperative magnetic resonance imaging (MRI). Groups 1 and 2 were similar in terms of age, body mass index, preoperative co‐morbidities (cardiovascular and diabetes mellitus), clinical stage of prostate cancer and preoperative PSA levels.
• ? In pathological specimens, low total testosterone (<3 ng/mL) was an independent risk factor for high Gleason score (>7) and for locally advanced pathological stage (pT3 and pT4).
• ? Higher preoperative testosterone correlated with disease confined to the gland.
• ? There was no association between serum testosterone levels and surgical margin status, on the one hand, and biochemical recurrence on the other.

CONCLUSION

• ? Low serum testosterone appears to be predictive of aggressive disease (Gleason score >7 and extraprostatic disease, pathological stage >pT2) in patients who underwent RP for localized prostate cancer.

     

Image-guided laser ablation of osteoid osteoma in pediatric patients

BACKGROUND: We aimed to evaluate the effectiveness of image-guided laser ablation for the treatment of osteoid osteomas in pediatric patients. METHODS: From June 1994 to June 2006, 68 patients aged 3 to 18 years (mean age, 12.1 +/- 4.2 years) with a diagnosis of osteoid osteoma underwent laser ablation in our department. Among them, 3 patients had prior unsuccessful attempt of surgical resection. Procedures were performed under spinal or general anesthesia with combined computed tomographic and fluoroscopic guidance. An 18-gauge needle was placed in the nidus after bone drilling when required, and an optical fiber was inserted through it. An average energy of 1200 J was delivered using an infrared diode laser generator. Patients were discharged the next day and followed up with assessment of pain, medication intakes, and potential complications. Clinical success was defined by complete pain relief without need for medication intake. RESULTS: Technical success was achieved in all but 1 of our first patients, which was subsequently treated by percutaneous resection. Overall clinical success was 98%. Primary clinical success was obtained in 60 patients (88%). In 7 patients, recurrence of symptoms at 4 to 27 months from the initial procedure was managed by repeat laser ablation, resulting in secondary success (10%). No complications related to the procedure were observed. Follow-up ranged 16 to 146 months (mean, 83 months). CONCLUSIONS: Currently, image-guided laser ablation is a safe and effective procedure that can be proposed as the mainstay treatment of osteoid osteomas in pediatrics. LEVEL OF EVIDENCE: Therapeutic case series study level IV.