Minimal Protocol Research Articles

Gonadotropin surge stimulate ovarian osteopontin expression via epidermal growth factor receptor signaling to enhance corpus luteum function

cific angiocrine factors, creating an instructive vascular niche which promotes organ development, repair and regeneration. Studying the role of primary ovarian ECs in the regulation of physiological processes in the ovary has been hampered by the need for growth factor deprivation during culture, which leads to apoptosis. Here, we aim to isolate, purify, characterize and culture ovarian ECs in serum-free conditions, creating a platform for the exploration of endothelial-follicular communication. DESIGN: Laboratory study. MATERIALS AND METHODS: Mouse ovarian ECs were isolated via magnetic cell sorting using CD-31 antibody. After a culture period of 5-7 days, EC activation was initiated via lentivirus transduction, allowing prolonged culture in serum-free conditions. Purity of cells was confirmed with flow cytometry using CD31, CD45 and VE-Cadherin antibodies (VECAD). Gene expression was assessed with qPCR. In order to obtain adequate numbers of fresh ECs for comparison, a minimal stimulation protocol was applied. Intravital staining using VECAD and Isolectin was performed, allowing ovarian EC isolation using florescence activated cell sorting (FACS). Gene expression of fresh ECs was assessed with qPCR and compared to cultured counterparts. RESULTS: Flow cytometry showed that 98% of cultured activated ovarian ECs were CD31+, VECAD+ and CD45-. Relative expression of dll1, mmp10 and fgf1 was higher in cultured ECs, while expression of Jagged-1, VECAD and PDGF was comparable in both groups. Expression of Jagged-2, Notch14, dll4, VEGFR1, VEGFR2, mmp4, il6, and il8 was lower in activated, cultured ECs in comparison to fresh ECs but fairly high compared to its own housekeeping gene. CONCLUSION: Here, we report successful isolation and durable culture of ovarian ECs with an angiocrine profile matching that of fresh ovarian ECs, thus providing a novel tool with which to study cell-cell communication and vascularization within the ovary. Supported by: Institutional.

A New Authentication Scheme for the Protection of FPGA Based IP Designs

Intellectual Property (IP) plays an essential part in the design-for-reuse technique which minimizes cost and development time of System-on-Chip (SoC) designs. But, sharing IP designs has many security risks and moreover, traditional methods for IP protection are time consuming and are often unaffordable and thus, protection of IP designs in VLSI have become an active research area. Field Programmable Gate Array (FPGA) uses bitstream encryption method to protect Intellectual Property (IP) cores once it is loaded onto the FPGA. Static Random Access Memory ( SRAM) based FPGA are volatile and the requirement of configuring on each power up results in attacks such as cloning, reverse engineering or tampering of the bitstream. So, advanced and novel techniques beyond bitstream encryption are necessary to ensure FPGA design security. This research work focuses on establishing an effective method for embedding IP designer information starting from synthesis tool level. High security is also provided to IP cores by using a novel hardware in the FPGA called Secure Start Hardware (SSH). A minimal cryptographic protocol is introduced to achieve an authenticated channel between the system designer (SD) and Secure Start Hardware (SSH). The proposed approach and design is tested and it is observed to provide efficient results.

A protocol for the treatment of retinopathy of prematurity in Spain

ObjectiveTo prepare a protocol for the treatment of retinopathy of prematurity (ROP) agreed by the majority of Spanish ophthalmologists dedicated to this topic. Material and methodA draft of the protocol was produced taking into account the experience of the participants and up to date publications. This draft was corrected by all the ophthalmologists participating in the project, and the final document was agreed by all of them. ResultsWe present general guidelines as an aid for the treatment of ROP, including treatment criteria, treatment methods, a calendar of action, and follow-up. ConclusionsIt is important to have a common working protocol for the treatment of ROP to improve care and to avoid mistakes. Although individual Hospitals may adapt the protocol to their daily activity, it is recommended that there is a minimal working protocol agreed by most of professionals dedicated to pediatric ophthalmology in Spain.

Screening program for retinopathy of prematurity in Spain

ObjectiveTo prepare a retinopathy of prematurity (ROP) screening program as agreed by most of Spanish ophthalmologists dedicated to this topic. Materials and methodsA draft of the protocol was produced taking into account the experience of the participants and current publications. This draft was corrected by all the ophthalmologists participating in the project and the final document produced was agreed by all of them. ResultsWe present general guidelines to help in the screening of ROP, including treatment criteria, treatment methods, and a calendar of action. ConclusionsIt is important to have a common working protocol in the screening of ROP to improve the action and to avoid mistakes. Although individual hospitals may adapt the protocol to their daily activity, it is recommended that there is a minimal working protocol agreed by most of professionals dedicated to pediatric ophthalmology in Spain.

Waste Minimization Protocols for the Process of Synthesizing Zeolites from South African Coal Fly Ash.

Production of a high value zeolite from fly ash has been shown to be an avenue for the utilization of South African fly ash which presently constitutes a huge disposal problem. The synthesis of zeolites Na-P1 and analcime on a micro-scale has been successful and preliminary investigation shows that scale-up synthesis is promising. However, the post-synthesis supernatant waste generated contains high levels of NaOH that may constitute a secondary disposal problem. A waste minimization protocol was developed to reduce the volume of waste generated with a view to enhancing the feasibility of the scale synthesis. Series of experiments were conducted in 100 mL jacketed batch reactors. Fly ash was reacted with 5 Mol NaOH on a 1:1 mass basis during the aging step, followed by hydrothermal treatment in which ultrapure water was added to the slurry. This study shows that by re-introducing the supernatant waste into the experiments in such a way that it supplies the required reagent (NaOH) for the zeolite synthesis, zeolite Na-P1 and analcime can be synthesized. It also shows that the synthesis process can be altered to allow up to 100% re-use of the supernatant waste to yield high value zeolitic products. This study effectively constructed two protocols for the minimization of waste generated during the synthesis of zeolites from South African coal fly ash. This result could be used to establish a basis for legal and environmental aspects involved in the commission of a full-scale plant synthesizing zeolites NaP1 and analcime.

Post‐stroke dysphagia: progress at last

Oropharyngeal Dysphagia (OD) is both underestimated and underdiagnosed as a cause of malnutrition and respiratory complications following stroke. OD occurs in more than 50% of stroke patients. Aspiration pneumonia (AP) occurs in up to 20% of acute stroke patients and is a major cause of mortality after discharge. Systematic screening for OD should be performed on every patient with stroke before starting oral feeding, followed, if appropriate by clinical and instrumental (videofluroscopy and/or fiberoptic endoscopy) assessment. Bolus modification with adaptation of texture and viscosity of solids and fluids and postural adjustments should be part of the minimal treatment protocol, but they do not change the impaired swallow physiology nor promote recovery of damaged neural swallow networks in stroke patients. To this purpose, two new neurostimulation approaches are being developed to stimulate cortical neuroplasticity to recover swallowing function: (i) those aimed at stimulating the peripheral oropharyngeal sensory system by chemical, physical or electrical stimulus; and (ii) those aimed at directly stimulating the pharyngeal motor cortex, such as repetitive transcranial magnetic stimulation (rTMS). The study of Park et al. in this issue of Neurogastroenterology and Motility evaluated the effect of rTMS in dysphagic stroke patients and showed a marked improvement in swallow physiology. Other studies also using rTMS showed plastic changes in pharyngeal motor cortical areas relevant to swallowing function. If further randomized controlled trials confirm these initial results, the neurorehabilitation strategies will be introduced to clinical practice sooner rather than later, improving the recovery of dysphagic stroke patients. Progress at last.

Calcineurin inhibitors in liver transplantation – still champions or threatened by serious competitors?

Current strategies for immunosuppression in liver transplant (LT) recipients include the design of protocols targeting a more individualized approach to reduce risk factors such as renal failure, cardiovascular complications and malignancies. Renal injury in LT recipients may be often multifactorial and is associated with increased risk of post-transplant morbidity and mortality. The quest for low toxicity immunosuppressive regimens has been challenging and resulted in CNI minimization protocols or CNI withdrawal and conversion to mycophenolate mofetil (MMF) and/or mammalian target of rapamycin inhibitor-based immunosuppressive regimens. Use of antibody induction to delay CNI administration may be an option in particular in immunocompromized, critically ill patients with high MELD scores. Protocols including MMF introduction and concomitant CNI minimization have the potential to recover renal function even in the medium and long term after LT. We review on hot topics in the prevention and management of acute and chronic renal injury in LT patients. For this purpose, we present and critically discuss results from immunosuppressive studies published in the current literature or presented at recent LT meetings.

Toward minimal conditioning protocols for allogeneic chimerism in tolerance resistant recipients

Mixed chimerism is a promising approach toward generating donor-specific immunological tolerance. However, chimerism induction can be toxic; therefore, there is an effort to develop non-myeloablative, minimal intensity protocols that can generate chimerism without the toxic side effects. Recently, with the goal of creating a minimalistic chimerism induction protocol in the tolerance resistant non-obese diabetic (NOD) mouse model, we identified pre-existing T cells as cells that resist fully allogeneic chimerism. With monoclonals targeting NOD T cells, we showed that long-term chimerism and tolerance toward donor islets could be established. However, this promising new protocol relied on the administration of a single dose of anti-CD40 ligand, which is not clinically applicable. In refining protocols to move even closer to clinical utility, we report here initial success at generating fully allogeneic mixed chimerism in NOD mice by adding cyclophosphamide to the conditioning regimen in place of anti-CD40 ligand antibodies.

Divergence in Defence against Herbivores between Males and Females of Dioecious Plant Species.

Defensive traits may evolve differently between sexes in dioecious plant species. Our current understanding of this process hinges on a partial view of the evolution of resistance traits that may result in male-biased herbivory in dioecious populations. Here, we present a critical summary of the current state of the knowledge of herbivory in dioecious species and propose alternative evolutionary scenarios that have been neglected. These scenarios consider the potential evolutionary and functional determinants of sexual dimorphism in patterns of resource allocation to reproduction, growth, and defence. We review the evidence upon which two previous reviews of sex-biased herbivory have concluded that male-biased herbivory is a rule for dioecious species, and we caution readers about a series of shortcomings of many of these studies. Lastly, we propose a minimal standard protocol that should be followed in any studies that intend to elucidate the (co)evolution of interactions between dioecious plants and their herbivores.

Flexible ovarian stimulation in a poor responder: a case report and literature review

This article reports on a novel strategy of continuous ovarian stimulation in a poor responder with two oocyte retrievals within the follicular and luteal phases of the same menstrual cycle. It also reviews studies of flexible ovarian stimulation. A patient aged 41years diagnosed as infertile with low ovarian reserve sought IVF treatment. A minimal ovarian simulation protocol was commenced with use of low-dose urinary FSH (150IU/day) from day 3, combined with using clomiphene citrate (50–100mg) when the biggest follicle reached 10mm in diameter to prevent ovulation. Gonadotrophin-releasing hormone agonist was administered on day 15 when the lead follicles reached 18mm in diameter. The first ultrasound-guided oocyte retrieval was performed 21h later; however, no ovum was obtained. Luteal-phase ovarian stimulation was started 2days later. The second oocyte retrieval was performed 25h after human chorionic gonadotrophin administration. Finally, one mature oocyte resulting in a cleavage embryo (8-cell, grade 2) was obtained. The case demonstrates that it is feasible to obtain mature oocytes and good embryos from luteal-phase stimulation. Continuous stimulation during the follicular and luteal phase can be a solution for poor ovarian response. Ovarian stimulation may be performed flexibly in special circumstances.This article reports on a novel strategy of continuous ovarian stimulation on a poor responder during the follicular and luteal phases of the same menstrual cycle with two oocyte retrievals. It also reviews studies of flexible ovarian stimulation. In our case, a patient, aged 41, who was diagnosed as infertile with very poor ovarian function, sought therapy for IVF. Antral follicles and basic hormones were evaluated on day 2 of the menstrual cycle. The stimulation cycle was commenced with use of 150IU/day urinary FSH from day 3, combined with clomiphene citrate and ibuprofen in the late follicular and luteal phases to prevent ovulation. Oocyte retrieval was attempted twice. We performed luteal-phase ovarian stimulation continuously 2days after the first failed oocyte retrieval during the follicular phase. Finally, one mature oocyte resulting in a cleavage embryo (8-cell, grade 2) was obtained after the luteal phase stimulation and retrieval, redeeming the previous unsuccessful oocyte retrieval. The case demonstrates that it is practical to obtain mature oocytes and good embryos from maintaining luteal-phase stimulation. For poor ovarian response, continuous stimulation during the follicular and luteal phases can be a solution. The literature review summarizes the clinical outcomes and parameters of flexible ovarian stimulation previously reported, including luteal-phase oocyte retrieval plus in-vitro maturation, ovarian stimulation initiated during the follicular or luteal phase and random-start stimulation. With knowledge gleaned from the case and review of studies, ovarian stimulation may be performed flexibly in special circumstances.

TEMPORAL HYGIENE: PROBLEMS IN CULTURAL CHRONOLOGY OF THE LATE PREHISTORIC PERIOD OF THE MIDDLE OHIO RIVER VALLEY

Radiocarbon dates are an independent source of data for constructing temporal relationships. Too often dates are shoehorned by preconceptions and hobbled by casual sample selection, reporting, and interpretation. Several authors have established quality-control criteria for evaluating and cleansing regional radiocarbon databases. Recent applications of such temporal hygiene practices in paleoecology and archaeology have resulted in significant reinterpretations of the sequence, timing, and tempo of change. Further, a new appreciation is often gained of the extent of synchronic variability. I apply a minimal hygienic protocol based on three criteria: material dated, method of age estimation, and when the sample was run. I illustrate both the value of and the need for such analyses by cleaning up a small sample of dates (N = 125) from 11 Fort Ancient sites. The single largest problem for determining occupation timing is the use of wood samples. Nearly 80 percent of the assays are unacceptable for use in reconstructing occupation history, except to frame the maximal age of occupation. The modifications to the timing and duration of occupation for these 11 sites significantly undermine the logical inferential structure of current reconstructions of change and difference in the middle Ohio River valley.

Lessons learned from war: a comprehensive review of the published experiences of the Iranian neurosurgeons during Iraq-Iran conflict and review of the related literature.

Background and Objective: The present study was aimed to review the articles published by Iranian neurosurgeons regarding their experiences during Iraq-Iran 8-year conflict and compare it with reports from other conflicts. Methods: The relevant papers and studies, published up to December 2011, were searched from three databases including MEDLINE, and two Iranian databases of IranMedex and Scientific Information Database with the keywords Iran, Iraq, conflict, battle, war, traumatic aneurysm (TA), post-traumatic epilepsy (PTE), brain infection, penetrating head wound (PHW), cerebrospinal fluid (CSF) leakage, spine injury and peripheral nerve injury. Results: A total of twenty-eight papers were found that presented PHW, development of TA, infections, PTE and peripheral nerve injuries. There were two different protocols for management of PHWs: radical surgery and minimal debridement protocol. The overall CNS infection rate was 10%. The cumulated incidence of TA was 6%. Conclusion: Conservative minimal debridement of the wounds was indicated in patients with small entrance wounds, or those with GCS≥ 8 and no progressive neurological deficit. To diagnose TA before rupture angiography was performed in patients with shells or bone fragments, passed through the crowded vasculature, or with large/delayed hematoma, or if the surgeon had high index of suspicion based on neuroimaging and early debridement surgery . Surgery in a well-equipped nearby hospital after quick and safe evacuation of the victims by trained-salvaging-ancillary groups, administration of broad-spectrum antibiotics and proper antiepileptic drugs decreased the morbidity and mortality of casualties after PHW in war situations. The biases of the case selection, data collection, confounders and decreasing biases by conducting blinded controlled clinical trials were discussed.__________________________________________________________ Full text of this presentation was published at the World Neurosurgery: http://www.worldneurosurgery.org/article/S1878-8750%2812%2900895-9/abstract

Minimal stimulation protocol using letrozole versus microdose flare up GnRH agonist protocol in women with poor ovarian response undergoing ICSI

Background: To compare the IVF outcomes of letrozole/antagonist and microdose GnRH agonist flare up protocols in poor ovarian responders undergoing intracytoplasmic sperm injection. Materials and methods: A randomized controlled trial was performed in patients with one or more previous failed IVF cycles in which four or less oocytes were retrieved when the gonadotrophin starting dose was at least 300 IU/day. Sixty patients were randomized by computer-generated list to receive either letrozole/antagonist (mild stimulation) n = 30 or GnRH-a protocol (microdose flare) n = 30. Results: Both groups were similar with respect to background and hormonal characteristics (age, duration of infertility, BMI, FSH, LH and E2). The clinical pregnancy rate per cycle was similar in both groups (13.3 vs. 16.6%; OR = 0.769; 95% CI = 0.185, 3.198). The doses of used gonadotropins and the number of stimulation days were significantly lower in the letrozole/antagonist protocol. The peak E2 level on the day of hCG, the endometrial thickness, the retrieved oocytes, the number of fertilized oocytes, the number of transferred embryos and the cancellation rate were statistically similar in both groups. Conclusions: The letrozole/antagonist protocol is a cost-effective and patient-friendly protocol that may be used in poor ovarian responders for IVF/ICSI.

The kidney in pediatric liver transplantation: An updated perspective

CKD continues to detract from the success of improved survival in pediatric liver transplantation, and its presence is likely under recognized. Here we review the literature regarding the prevalence, etiology, and management of renal dysfunction in pediatric liver transplant recipients. Long-term studies suggest the prevalence of CKD to be 25-38% by 5-10 yr post-transplant. While important, sole use of serum creatinine overestimates renal function in this population. Screening for and treatment of persistent proteinuria and hypertension as well as minimization of nephrotoxic insults are the mainstays to delay or prevent CKD progression. Office-based blood pressure measures are less sensitive than ABPM, which is specifically recommended by the American Heart Association for its ability to diagnose masked hypertension in pediatric liver transplant recipients. Long-term risk of CKD is predominantly secondary to CNI toxicity. CNI minimization protocols have shown promise in slowing progression of CKD while maintaining graft function, but large-scale randomized control trials with long-term follow-up are needed.

Renal Function Improvement After Conversion to Proliferation Signal Inhibitors During Long-Term Follow-up in Heart Transplant Recipients

IntroductionThe use of proliferation signal inhibitors (PSIs) for calcineurin-inhibitor (CNI) minimization or conversion protocols has been promoted for heart transplantation (HT) in the contexts of renal insufficiency, cardiac allograft vasculopathy (CAV), or malignancy. We evaluated our experience with conversion of patients from a CNI-based to a PSI-based immunosuppressive regimen. We focused on improvement in renal function. MethodsThis prospective follow-up included 96 HT patients converted to a PSI-based regimen from 2001 to 2010. We evaluated changes in creatinine clearance (CrCl) prior to at 1 year and at the end of follow-up after conversion. ResultsNinety-six patients including 86% men showed a mean age of 62 ± 8 years. They were converted to a PSI-based regimen at 6.3 ± 4 years post-HT due to the following causes: CNI toxicity (45%), CAV (16%), cancer (16%), CNI toxicity + CAV (17%), or CNI toxicity + cancer (6%). CNI withdrawal was achieved in 77 cases (80%) and minimization in 19 (20%). Everolimus was used in 54 (56%) and sirolimus in 42 (44%) cases. Median follow-up time was 3.8 years. PSI discontinuation due to side effects was common (38%). There were 43 deaths mainly due to cancer and CAV. CrCl improved albeit not significantly in the withdrawal group from a median of 51 mL/min preconversion to 59 mL/min at the last follow-up (P = .12). In the minimization group, median CrCl worsened from a median of 61 mL/min preconversion to 51 mL/min at the last follow-up (P = .001). In the 58 cases (61%) of CNI nephrotoxicity, median CrCl improved from a median of 41 mL/min preconversion to 49 mL/min at the last follow-up (P = .04). ConclusionDespite high rates of discontinuation of PSIs during long-term follow-up, the conversion regimen seemed to be useful to diminish CNI-related renal insufficiency especially with CNI withdrawal.

Protocolo de tratamiento de la retinopatía del prematuro en España

ObjectiveTo prepare a protocol for the treatment of retinopathy of prematurity (ROP) agreed by the majority of Spanish ophthalmologists dedicated to this topic. Material and methodA draft of the protocol was produced taking into account the experience of the participants and up to date publications. This draft was corrected by all the ophthalmologists participating in the project, and the final document was agreed by all of them. ResultsWe present general guidelines as an aid for the treatment of ROP, including treatment criteria, treatment methods, a calendar of action, and follow-up. ConclusionsIt is important to have a common working protocol for the treatment of ROP to improve care and to avoid mistakes. Although individual Hospitals may adapt the protocol to their daily activity, it is recommended that there is a minimal working protocol agreed by most of professionals dedicated to pediatric ophthalmology in Spain.

Programa de cribado para la retinopatía del prematuro en España

ObjectiveTo prepare a retinopathy of prematurity (ROP) screening program as agreed by most of Spanish ophthalmologists dedicated to this topic. Material and methodA draft of the protocol was produced taking into account the experience of the participants and current publications. This draft was corrected by all the ophthalmologists participating in the project and the final document produced was agreed by all of them. ResultsWe present general guidelines to help in the screening of ROP, including treatment criteria, treatment methods, and a calendar of action. ConclusionsIt is important to have a common working protocol in the screening of ROP to improve the action and to avoid mistakes. Although individual Hospitals may adapt the protocol to their daily activity, it is recommended that there is a minimal working protocol agreed by most of professionals dedicated to pediatric ophthalmology in Spain.

Juvenile mossy fibres: two pipers in the same pub?

Juvenile mossy fibres: two pipers in the same pub?

Recent Advances in Assisted Reproductive Technology

In 1978 the world witnessed the birth of the first “test tube baby.” Since this time there have been explosive advances in assisted reproductive technologies (ART). Current optimizations surrounding the delivery of in vitro fertilization (IVF) including the utilization of minimal stimulation protocols and gonadotropin releasing hormone (GnRH) agonist cycle triggers are being increasingly utilized to maximize patient safety. Modifications, such as those seen in the embryology laboratory, continue to improve pregnancy rates. Concurrent with these advancements in IVF have been the emergence of related technologies, such as embryonic genetic testing and oocyte preservation, which potentially have broad applications to both fertile and infertile couples. As these relevant applications of ART become increasingly utilized, it is incumbent upon society to ensure that these resources are made available in a morally responsible and equitable manner.

Crystal Growth, Structure and Morphology of Rifapentine Methanol Solvate

Rifapentine, an important antibiotic, was crystallized from methanol solvent in the form of its methanol solvate. The crystal structure of rifapentine methanol solvate belongs to monoclinic, space group P21, with the unit cell parameters of a = 1.2278(3) nm, b = 1.9768(4) nm, c = 1.2473(3) nm, Z = 2, and β = 112.35(3)°. The parallelepiped morphology was also predicted by Materials Studio simulation program. The influence of intermolecular interaction was taken into account in the attachment energy model. The crystal shape fits the calculated morphology well, which was performed on the potential energy minimized model using a generic DREIDING 2.21 force field and developed minimization protocol with derived partial charges.