Mosaic Turner Variant Adult Female Presenting with XO/XY Karyotype.

Turner syndrome (TS) is the most frequently detected chromosomal abnormality in females caused by the partial or complete absence of second X chromosome. Due to varied phenotypical presentation, the diagnosis of TS can create a spectrum of clinical concerns related to morbidity and mortality. At least 10% of Turner females exhibit the presence of Y chromosome or Y-derived sequences. Patients with 45,X/46,XY mosaicism may have a phenotypic variation of the external genitalia and exhibit features ranging from normal male to ambiguous to female genitalia with features of TS. Turner mosaic variants with Y chromosome components have increased risk for gonadoblastoma. Although the risk is not exactly quantifiable, according to the 2016 Cincinnati International TS Meeting Clinical Practice guidelines, bilateral prophylactic gonadectomy is mandatory if Y chromosomal component is identified in mosaic Turner. We describe a rare case of an adult female patient detected as mosaic Turner variant with the presence of Y chromosome and reconfirmed by an aneuploidy FISH probe.

Unravelling the Impact of an Additional Sex Chromosome in an Adult Female.

Women with Triple X syndrome (TXS) appear to be at increased risk for decreased ovarian reserve; however, available data are limited. We present an asyndromic adult female with features of recurrent pregnancy loss and decreased ovarian reserve detected with mosaic Triple X syndrome (TXS). The patient was initially evaluated by a low-cost peripheral blood (PB) conventional karyotyping using standard cytogenetic protocols. Interphase fluorescence in situ hybridisation was performed to confirm the diagnosis. Chromosomal microarray, which is a more expensive test, substantiated the presence of additional X chromosomes but failed to detect the presence of low level of mosaicism. Our case study emphasised the recommendation of performing a strategy-based cost-effective cytogenetic evaluation of all cases of decreased ovarian reserve or low anti-Müllerian hormone levels in a resource-constrained setting. It also highlighted the need for additional research to understand the natural history of ovarian function in TXS affected women throughout their lifespans.

Prevalence of Polycystic Ovarian Syndrome, Phenotypes and their Ovulation Response to Sequential Letrozole Dose Escalation among Infertile Women at a Tertiary Care Centre in Southern India.

Background: Women with polycystic ovarian syndrome (PCOS) often have anovulatory infertility requiring ovulation induction with letrozole. Aims: This study aimed to determine the prevalence and phenotypic categorisation of infertile PCOS women and to assess ovulatory response and pregnancy rates of PCOS phenotypes with sequential letrozole dose escalation. Study Setting and Design: This was a prospective observational study. Materials and Methods: One hundred seventy-five infertile PCOS women were enrolled. One hundred fifty-six women received ovulation induction as per the protocol with sequential letrozole dose escalation in each subsequent cycle (2.5 mg, 5 mg and 7.5 mg). Responses were assessed by ovulation and/or pregnancy. Statistical Analysis Used: Descriptive statistics were elaborated by means, medians, frequencies and percentages. Group comparisons and linear correlation between two continuous variables were done using appropriate statistical tests. Results: Eighty-seven (49.7%) women were Phenotype A; 11 (6.3%) were Phenotype B; 20 (11.4%) were Phenotype C and 57 (32.6%) were Phenotype D in our study. After excluding the lost to follow up participants in each induction cycle, 33.3% (2.5 mg dose); 62.8% (5 mg dose) and 78.9% (7.5 mg dose) women responded to letrozole. A significant increase in ovulation to escalating letrozole doses was noted (Phenotype A: 35.1% to 2.5 mg, 53.7% to 5 mg and 72.7% to 7.5 mg; Phenotype B: 30% to 2.5 mg and 80% to 5 mg; Phenotype C: 35.3% to 2.5 mg and 87.5% to 5 mg and Phenotype D: 30.8% to 2.5 mg, 65.6% to 5 mg and 87.5% to 7.5 mg). Fifty-six of 156 (35.9%) infertile PCOS women achieved pregnancy; increase in pregnancy rates with escalated doses of letrozole was noted. Conclusion: All PCOS phenotypes show a similar response to escalating doses of letrozole. The role of phenotypic sub-categorisation for variable response to letrozole as an ovulation-inducing agent is uncertain.

Characteristics of Empty Follicular Syndrome during In vitro Fertilization Embryo Transfer and its Association with Various Etiologies in Comparatively Young Patients.

BACKGROUND: Nearly 0.6%-7% of patients undergoing in vitro fertilization embryo transfer (IVF ET) will not be able to yield any oocyte despite successful ovarian stimulation and this condition is called as empty follicular syndrome (EFS). EFS is a dreadful situation for clinicians as well as patients, seems to be an unavoidable clinical condition despite a proper ovarian stimulation. MATERIALS AND METHODS: This was a retrospective observational study conducted at a tertiary hospital; 1103 patients who underwent IVF ET between January 2016 and May 2017 were included in the study. STUDY OUTCOME: To estimate the incidence of empty follicle syndrome (EFS) and to study the associated factors. RESULTS: There were 53 (4.8%) cases of EFS out of 1103 cycles of IVF ET; 43 (3.9%) cases were false EFS and 10 (0.9%) cases were genuine EFS. Mean age of EFS group and oocyte retrieved group was 30.17 years and 29.12 years respectively. Recurrence rate of EFS during the next IVF cycle was 36.8%. Decreased ovarian reserve was associated with an increased chance of EFS (54.7%) with a recurrence rate as high as 57%. CONCLUSION: The incidence of EFS is not an uncommon clinical scenario; it depends upon ovarian reserve to a great extent. Young age is not immune for the occurrence of EFS as there is a similar incidence in comparatively younger age group in our study. EFS is seen in all etiological groups of infertility, but only respite is that there is a chance of about 63.2% oocyte retrieval during repeat IVF cycle.

Correlation between Serum Progesterone Level on the Day of Ovulation Trigger During In vitro Fertilization and Its Effect on Treatment Outcome.

BACKGROUND: Premature luteinization (PL) is defined as a premature rise in serum progesterone concentration on or before the day of ovulation trigger with human chorionic gonadotropin. The incidence of PL varies between 5% and 30% during in vitro fertilization and embryo transfer (IVF-ET). MATERIALS AND METHODS: The prospective observational study comprising 380 patients undergoing IVF-ET. Blood samples were collected for serum progesterone level estimation on the day of ovulation trigger. Ovum pickup was done 36 h later and serum progesterone levels were correlated with IVF-ET outcome. STUDY OUTCOME: To correlate serum progesterone level on the day of ovulation trigger during IVF and its effect on treatment outcome. RESULTS: Mean serum progesterone level in the positive pregnancy group and negative pregnancy group was 0.892 ± 0.752 ng/ml and 0.91 ± 0.688 ng/ml, respectively (P = 0.961). The overall incidence of PL was 12.8% with 12.7% and 13.6% in the agonist and antagonist protocol respectively (P = 0.9001). PL incidence was 13.5% and 13.4% in positive pregnancy and negative pregnancy group (P = 0.223). CONCLUSION: PL has been associated with 12.8% of the IVF cycles. There was no statistically significant difference observed in the incidence of PL between different IVF stimulation protocols. PL does not seem to affect IVF outcome.

A study on cervical cancer screening in asymptomatic women using Papanicolaou smear in a tertiary care hospital in an urban area of Mumbai, India.

INTRODUCTION: Cervical cancer is the most common cause of death among women in developing countries. Among the Indian women, cervical cancer is the most common genital tract cancer. Papanicolaou (Pap) smear test plays a vital role in the detection of cervical cancer even in its premalignant condition. The aim of this study to evaluate the role of Pap smear in detecting premalignant and malignant lesions as well as nonneoplastic lesions of the cervix and to determine the prevalence of various lesions. MATERIALS AND METHODS: We screened 1100 women in the age group of 21-65 years who attended our medical camp organized by the hospital in outdoor patient department. All women was willing to give consent for screening by Pap smear test were included. RESULTS: Of 1100 cases, majority of the cases were benign comprising negative for intraepithelial neoplasia (NILM) of about 581 (52.8%) cases, 203 (18.4%) inflammatory, atypical squamous cells of undetermined significance 45 (4%), low-grade squamous intraepithelial lesion (LSIL) in 75 (6.8%), and high-grade squamous intraepithelial lesion (HSIL) in 74 (6%) women. Overall sensitivity and specificity for the detection of LSIL were 75.8% and 94.6% and those for the detection of HSIL were 68.9% and 98.6%. CONCLUSIONS: Pap smear test is a very easy, noninvasive, useful, simple, safe, and very economical tool to detect preinvasive cervical epithelial lesions. It is evident and proven that every woman above the age of 30-35 years must be subjected to cervical screening and this must be continued even in the postmenopausal period.

CB1R-Mediated Activation of Caspase-3 Causes Epigenetic and Neurobehavioral Abnormalities in Postnatal Ethanol-Exposed Mice.

Alcohol exposure can affect brain development, leading to long-lasting behavioral problems, including cognitive impairment, which together is defined as fetal alcohol spectrum disorder (FASD). However, the fundamental mechanisms through which this occurs are largely unknown. In this study, we report that the exposure of postnatal day 7 (P7) mice to ethanol activates caspase-3 via cannabinoid receptor type-1 (CB1R) in neonatal mice and causes a reduction in methylated DNA binding protein (MeCP2) levels. The developmental expression of MeCP2 in mice is closely correlated with synaptogenesis and neuronal maturation. It was shown that ethanol treatment of P7 mice enhanced Mecp2 mRNA levels but reduced protein levels. The genetic deletion of CB1R prevented, and administration of a CB1R antagonist before ethanol treatment of P7 mice inhibited caspase-3 activation. Additionally, it reversed the loss of MeCP2 protein, cAMP response element binding protein (CREB) activation, and activity-regulated cytoskeleton-associated protein (Arc) expression. The inhibition of caspase-3 activity prior to ethanol administration prevented ethanol-induced loss of MeCP2, CREB activation, epigenetic regulation of Arc expression, long-term potentiation (LTP), spatial memory deficits and activity-dependent impairment of several signaling molecules, including MeCP2, in adult mice. Collectively, these results reveal that the ethanol-induced CB1R-mediated activation of caspase-3 degrades the MeCP2 protein in the P7 mouse brain and causes long-lasting neurobehavioral deficits in adult mice. This CB1R-mediated instability of MeCP2 during active synaptic maturation may disrupt synaptic circuit maturation and lead to neurobehavioral abnormalities, as observed in this animal model of FASD.

Innominate Vein Stenosis in Association with Ipsilateral Hyperdynamic Brachiobasilic Fistula Causing Ipsilateral Limb and Hemifacial Swelling.

A 34-year-old hypertensive woman with a hyperdynamic, left brachiobasilic dialysis fistula presented with a long history of throbbing in her head and swelling of the left side of the face. Tight stenosis of left brachiocephalic vein was found to be causing retrograde flow into the left jugular vein which normalized after dilatation and stenting with resolution of all the symptoms and patient is asymptomatic for 1 year.

Diagnostic Office Vaginohysteroscopy in Evaluation of Infertility Prior to IVF: A Retrospective Analysis of 1000 Cases.

AIM: The aim of this study was to analyze the utility of routine use of diagnostic office vaginohysteroscopy in the evaluation of uterine cavity in infertility patients prior to IVF-ET. MATERIALS AND METHODS: We conducted a retrospective analysis of 1000 women who had undergone routine diagnostic office vaginohysteroscopy as an institutional protocol in the evaluation of infertility prior to IVF-ET cycle at a tertiary care hospital. They were divided into two groups: primary infertility (group I) and secondary infertility (group II). The primary outcome was the finding of an abnormal uterine cavity (congenital abnormality vs acquired abnormality). RESULTS: One thousand women underwent routine diagnostic office vaginohysteroscopy in the evaluation of infertility prior to IVF-ET. There were no intraoperative or postoperative complications. Vaginohysteroscopy revealed an abnormal uterine cavity in 13.8% (1000 patients) of women. Primary infertility group (I) had 13.19% (811 patients), and secondary infertility group (II) had 16.4% (189 patients) abnormal uterine cavities. CONCLUSION: Diagnostic office vaginohysteroscopy has a definite role in the uterine cavity evaluation in infertility patients prior to IVF, but routine use should not be recommended considering the low incidence of abnormal uterine cavity findings. Moreover, the majority of these uterine cavity abnormalities can be detected by less invasive tests such as HSG, TVS, SSG and 3D ultrasound.

A single day of 5-azacytidine exposure during development induces neurodegeneration in neonatal mice and neurobehavioral deficits in adult mice.

The present study was undertaken to evaluate the immediate and long-term effects of a single-day exposure to 5-Azacytidine (5-AzaC), a DNA methyltransferase inhibitor, on neurobehavioral abnormalities in mice. Our findings suggest that the 5-AzaC treatment significantly inhibited DNA methylation, impaired extracellular signal-regulated kinase (ERK1/2) activation and reduced expression of the activity-regulated cytoskeleton-associated protein (Arc). These events lead to the activation of caspase-3 (a marker for neurodegeneration) in several brain regions, including the hippocampus and cortex, two brain areas that are essential for memory formation and memory storage, respectively. 5-AzaC treatment of P7 mice induced significant deficits in spatial memory, social recognition, and object memory in adult mice and deficits in long-term potentiation (LTP) in adult hippocampal slices. Together, these data demonstrate that the inhibition of DNA methylation by 5-AzaC treatment in P7 mice causes neurodegeneration and impairs ERK1/2 activation and Arc protein expression in neonatal mice and induces behavioral abnormalities in adult mice. DNA methylation-mediated mechanisms appear to be necessary for the proper maturation of synaptic circuits during development, and disruption of this process by 5-AzaC could lead to abnormal cognitive function.

An investigation into the role of ATP in the mammalian pre-mRNA 3' cleavage reaction.

RNA Polymerase II transcribes beyond what later becomes the 3' end of a mature messenger RNA (mRNA). The formation of most mRNA 3' ends results from pre-mRNA cleavage followed by polyadenylation. In vitro studies have shown that low concentrations of ATP stimulate the 3' cleavage reaction while high concentrations inhibit it, but the origin of these ATP effects is unknown. ATP might enable a cleavage factor kinase or activate a cleavage factor directly. To distinguish between these possibilities, we tested several ATP structural analogs in a pre-mRNA 3' cleavage reaction reconstituted from DEAE-fractionated cleavage factors. We found that adenosine 5'-(ß,γ-methylene)triphosphate (AMP-PCP) is an effective in vitro 3' cleavage inhibitor with an IC50 of ∼300 µM, but that most other ATP analogs, including adenosine 5'-(ß,γ-imido)triphosphate, which cannot serve as a protein kinase substrate, promoted 3' cleavage but less efficiently than ATP. In combination with previous literature data, our results do not support ATP stimulation of 3' cleavage through cleavage factor phosphorylation in vitro. Instead, the more likely mechanism is that ATP stimulates cleavage factor activity through direct cleavage factor binding. The mammalian 3' cleavage factors known to bind ATP include the cleavage factor II (CF IIm) Clp1 subunit, the CF Im25 subunit and poly(A) polymerase alpha (PAP). The yeast homolog of the CF IIm complex also binds ATP through yClp1. To investigate the mammalian complex, we used a cell-line expressing FLAG-tagged Clp1 to co-immunoprecipitate Pcf11 as a function of ATP concentration. FLAG-Clp1 co-precipitated Pcf11 with or without ATP and the complex was not affected by AMP-PCP. Diadenosine tetraphosphate (Ap4A), an ATP analog that binds the Nudix domain of the CF Im25 subunit with higher affinity than ATP, neither stimulated 3' cleavage in place of ATP nor antagonized ATP-stimulated 3' cleavage. The ATP-binding site of PAP was disrupted by site directed mutagenesis but a reconstituted 3' cleavage reaction containing a mutant PAP unable to bind ATP nevertheless underwent ATP-stimulated 3' cleavage. Fluctuating ATP levels might contribute to the regulation of pre-mRNA 3' cleavage, but the three subunits investigated here do not appear to be responsible for the ATP-stimulation of pre-mRNA cleavage.

CB1-receptor knockout neonatal mice are protected against ethanol-induced impairments of DNMT1, DNMT3A, and DNA methylation.

The significant consequences of ethanol use during pregnancy are neurobehavioral abnormalities involving hippocampal and neocortex malfunctions that cause learning and memory deficits collectively named fetal alcohol spectrum disorder. However, the molecular mechanisms underlying these abnormalities are still poorly understood and therefore warrant systematic research. Here, we document novel epigenetic abnormalities in the mouse model of fetal alcohol spectrum disorder. Ethanol treatment of P7 mice, which induces activation of caspase 3, impaired DNA methylation through reduced DNA methyltransferases (DNMT1 and DNMT3A) levels. Inhibition of caspase 3 activity, before ethanol treatment, rescued DNMT1, DNMT3A proteins as well as DNA methylation levels. Blockade of histone methyltransferase (G9a) activity or cannabinoid receptor type-1 (CB1R), prior to ethanol treatment, which, respectively, inhibits or prevents activation of caspase 3, rescued the DNMT1 and DNMT3A proteins and DNA methylation. No reduction of DNMT1 and DNMT3A proteins and DNA methylation was found in P7 CB1R null mice, which exhibit no ethanol-induced activation of caspase 3. Together, these data demonstrate that ethanol-induced activation of caspase 3 impairs DNA methylation through DNMT1 and DNMT3A in the neonatal mouse brain, and such impairments are absent in CB1R null mice. Epigenetic events mediated by DNA methylation may be one of the essential mechanisms of ethanol teratogenesis. Schematic mechanism of action by which ethanol impairs DNA methylation. Studies have demonstrated that ethanol has the capacity to bring epigenetic changes to contribute to the development of fetal alcohol spectrum disorder (FASD). However, the mechanisms are not well studied. P7 ethanol induces the activation of caspase 3 and impairs DNA methylation through reduced DNA methyltransferases (DNMT1 and DNMT3A) proteins (→). The inhibition or genetic ablation of cannabinoid receptor type-1 or inhibition of histone methyltransferase (G9a) by Bix (-----) or inhibition of caspase 3 activation by Q- quinoline-Val-Asp(Ome)-CH2-O-phenoxy (Q-VD-OPh) () rescue loss of DNMT1, DNMT3A as well as DNA methylation. Hence, the putative DNMT1/DNMT3A/DNA methylation mechanism may have a potential regulatory role in FASD.

Elevation of endogenous anandamide impairs LTP, learning, and memory through CB1 receptor signaling in mice.

In rodents, many exogenous and endogenous cannabinoids, such as anandamide (AEA) and 2-arachidonyl glycerol (2-AG), have been shown to play an important role in certain hippocampal memory processes. However, the mechanisms by which endogenous AEA regulate this processes are not well understood. Here the effects of AEA on long-term potentiation (LTP), hippocampal-dependent learning and memory tasks, pERK1/2, pCaMKIV, and pCREB signaling events in both cannabinoid receptor type 1 (CB1R) wild-type (WT) and knockout (KO) mice were assessed following administration of URB597, an inhibitor of the fatty acid amide hydrolase (FAAH). Acute administration of URB597 enhanced AEA levels without affecting the levels of 2-AG or CB1R in the hippocampus and neocortex as compared to vehicle. In hippocampal slices, URB597 impaired LTP in CB1R WT but not in KO littermates. URB597 impaired object recognition, spontaneous alternation and spatial memory in the Y-maze test in CB1R WT mice but not in KO mice. Furthermore, URB597 enhanced ERK phosphorylation in WT without affecting total ERK levels in WT or KO mice. URB597 impaired CaMKIV and CREB phosphorylation in WT but not in KO mice. CB1R KO mice have a lower pCaMKIV/CaMKIV ratio and higher pCREB/CREB ratio as compared to WT littermates. Our results indicate that pharmacologically elevated AEA impair LTP, learning and memory and inhibit CaMKIV and CREB phosphorylation, via the activation of CB1Rs. Collectively, these findings also suggest that pharmacological elevation of AEA beyond normal concentrations is also detrimental for the underlying physiological responses.

Ethanol exposure induces neonatal neurodegeneration by enhancing CB1R Exon1 histone H4K8 acetylation and up-regulating CB1R function causing neurobehavioral abnormalities in adult mice.

BACKGROUND: Ethanol exposure to rodents during postnatal day 7 (P7), which is comparable to the third trimester of human pregnancy, induces long-term potentiation and memory deficits. However, the molecular mechanisms underlying these deficits are still poorly understood. METHODS: In the present study, we explored the potential role of epigenetic changes at cannabinoid type 1 (CB1R) exon1 and additional CB1R functions, which could promote memory deficits in animal models of fetal alcohol spectrum disorder. RESULTS: We found that ethanol treatment of P7 mice enhances acetylation of H4 on lysine 8 (H4K8ace) at CB1R exon1, CB1R binding as well as the CB1R agonist-stimulated GTPγS binding in the hippocampus and neocortex, two brain regions that are vulnerable to ethanol at P7 and are important for memory formation and storage, respectively. We also found that ethanol inhibits cyclic adenosine monophosphate response element-binding protein (CREB) phosphorylation and activity-regulated cytoskeleton-associated protein (Arc) expression in neonatal and adult mice. The blockade or genetic deletion of CB1Rs prior to ethanol treatment at P7 rescued CREB phosphorylation and Arc expression. CB1R knockout mice exhibited neither ethanol-induced neurodegeneration nor inhibition of CREB phosphorylation or Arc expression. However, both neonatal and adult mice did exhibit enhanced CREB phosphorylation and Arc protein expression. P7 ethanol-treated adult mice exhibited impaired spatial and social recognition memory, which were prevented by the pharmacological blockade or deletion of CB1Rs at P7. CONCLUSIONS: Together, these findings suggest that P7 ethanol treatment induces CB1R expression through epigenetic modification of the CB1R gene, and that the enhanced CB1R function induces pCREB, Arc, spatial, and social memory deficits in adult mice.

Structurally diverse low molecular weight activators of the mammalian pre-mRNA 3' cleavage reaction.

The 3' end formation of mammalian pre-mRNA contributes to gene expression regulation by setting the downstream boundary of the 3' untranslated region, which in many genes carries regulatory sequences. A large number of protein cleavage factors participate in this pre-mRNA processing step, but chemical tools to manipulate this process are lacking. Guided by a hypothesis that a PPM1 family phosphatase negatively regulates the 3' cleavage reaction, we have found a variety of new small molecule activators of the in vitro reconstituted pre-mRNA 3' cleavage reaction. New activators include a cyclic peptide PPM1D inhibitor, a dipeptide with modifications common to histone tails, abscisic acid and an improved l-arginine ß-naphthylamide analog. The minimal concentration required for in vitro cleavage has been improved from 200µM to the 200nM-100µM range. These compounds provide unexpected leads in the search for small molecule tools able to affect pre-mRNA 3' end formation.

A quantitative comparison between free uroflow variables and urodynamic data, and the effect of the size of urodynamic catheters on its interpretation.

OBJECTIVE: To assess the effect of the urodynamic catheter on the urinary flow rate and residual volume in various urodynamic diagnoses, and compare the outcome when using a smaller catheter, as the effect of this catheter on free uroflow variables is mostly studied in patients with bladder outlet obstruction (BOO) and little is known about its effect in other urodynamic diagnoses. PATIENTS AND METHODS: In all, 319 men undergoing a pressure-flow study (PFS) with a 5 F filling and 5 F measuring bladder catheter were subdivided into three groups based on a urodynamic diagnosis, i.e. normal PFS (group 1), BOO (group 2) and detrusor underactivity (DU, group 3). Another group (4) comprised 61 patients who had a PFS with the filling catheter removed before the voiding phase. The effect of the catheters on the maximum urinary flow rate (Qmax) and the postvoid residual volume (PVR) was analysed statistically and compared among the groups. We also compared the free-flow variables with the clinical and urodynamic variables. RESULTS: Groups 1-3 (with two catheters) had a significantly lower Qmax and higher PVR than those voiding with one catheter (group 4). The reduction in Qmax was highest in group 3 (41.9%) and least in group 2 (21%). Group 4 showed no significant change in Qmax in cases with BOO and a normal PFS but a significant decline in those with DU (19.6%). The PVR was positively associated with the bladder capacity and negatively with detrusor contractility, but no association with a urodynamic diagnosis of BOO or any specific symptom. CONCLUSION: Detrusor contractility was the strongest predictor of the obstructive effect caused by the catheter. This study justifies the use of a single 5 F catheter at the time of voiding, although that can also cause a reduction in flow in patients with DU.

The TF-antigen binding lectin from Sclerotium rolfsii inhibits growth of human colon cancer cells by inducing apoptosis in vitro and suppresses tumor growth in vivo.

Glycan array analysis of Sclerotium rolfsii lectin (SRL) revealed its exquisite binding specificity to the oncofetal Thomsen-Friedenreich (Galß1-3GalNAcα-O-Ser/Thr, T or TF) antigen and its derivatives. This study shows that SRL strongly inhibits the growth of human colon cancer HT29 and DLD-1 cells by binding to cell surface glycans and induction of apoptosis through both the caspase-8 and -9 mediated signaling. SRL showed no or very weak binding to normal human colon tissues but strong binding to cancerous and metastatic tissues. Intratumor injection of SRL at subtoxic concentrations in NOD-SCID mice bearing HT29 xenografts resulted in total tumor regression in 9 days and no subsequent tumor recurrence. As the increased expression of TF-associated glycans is commonly seen in human cancers, SRL has the potential to be developed as a therapeutic agent for cancer.

CD45-mediated signaling pathway is involved in Rhizoctonia bataticola lectin (RBL)-induced proliferation and Th1/Th2 cytokine secretion in human PBMC.

We earlier reported the mitogenic and immunostimulatory activities of Rhizoctonia bataticola lectin (RBL), purified from phytopathogenic fungus R. bataticola in human PBMC. The lectin demonstrates specificity towards glycoproteins containing complex N-glycans. Since CD45-protein tyrosine phosphatase that abundantly expresses N-glycans is important in T-cell signaling, the study aimed to investigate the involvement of CD45 in the immunomodulatory activities of RBL. Flowcytometry and confocal microscopy studies revealed that RBL exhibited binding to PBMC and colocalized with CD45. The binding was comparable in cells expressing different CD45 isoforms-RA, -RB and -RO. CD45 blocking antibody reduced the binding and proliferation of PBMC induced by RBL. CD45-PTPase inhibitor dephostatin inhibited RBL-induced proliferation, expression of CD25 and pZAP-70. RBL-induced secretion of Th1/Th2 cytokines were significantly inhibited in presence of dephostatin. Also, dephostatin blocked phosphorylation of p38MAPK and STAT-5 that was crucial for the biological functions of RBL. The study demonstrates the involvement of CD45-mediated signaling in RBL-induced PBMC proliferation and Th1/Th2 cytokine secretion through activation of p38MAPK and STAT-5.

Purification and characterization of a mitogenic lectin from cephalosporium, a pathogenic fungus causing mycotic keratitis.

Ophthalmic mycoses caused by infectious fungi are being recognized as a serious concern since they lead to total blindness. Cephalosporium is one amongst several opportunistic fungal species implicated in ophthalmic infections leading to mycotic keratitis. A mitogenic lectin has been purified from the mycelia of fungus Cephalosporium, isolated from the corneal smears of a keratitis patient. Cephalosporium lectin (CSL) is a tetramer with subunit mass of 14 kDa, agglutinates human A, B, and O erythrocytes, and exhibits high affinity for mucin compared to fetuin and asialofetuin but does not bind to simple sugars indicating its complex sugar specificity. CSL showed strong binding to normal human peripheral blood mononuclear cells (PBMCs) to elicit mitogenic activity. The sugar specificity of the lectin and its interaction with PBMCs to exhibit mitogenic effect indicate its possible role in adhesion and infection process of Cephalosporium.

Rhizoctonia bataticola lectin (RBL) induces mitogenesis and cytokine production in human PBMC via p38 MAPK and STAT-5 signaling pathways.

BACKGROUND: Rhizoctonia bataticola lectin (RBL), purified from phytopathogenic fungus Rhizoctonia bataticola is highly mitogenic towards human peripheral blood mononuclear cells (PBMC). The lectin has sugar specificity towards N-glycans and binds to glycoproteins containing complex N-glycans (Nagre et al., Glycoconj J. 2010). In this study, we investigated the role of Mitogen Activated Protein Kinase (MAPK) and Signal Transducers and Activators of Transcription (STAT)-5 signaling in RBL-induced proliferation and production of Th1/Th2 cytokines. METHODS: Human PBMC were stimulated with RBL and proliferation was determined by tritiated thymidine incorporation assay, cytokine profiles by ELISA and activation of MAPK and STAT-5 by western blotting. RBL binding was monitored by immunofluorescence staining. Expression of IL-2Rα (CD25) was measured by flow cytometry. RESULTS: The binding and mitogenic activities of RBL were inhibited by glycoproteins- mucin, asialofetuin and fetuin. RBL stimulated expression of IL-2Rα and production of Th1/Th2 cytokines- IL-2, IFN-γ, IL-4 and IL-10. RBL-induced phosphorylation of ERK1/2 and p38 MAPK was detected at 1h and 3h respectively. Significant phosphorylation of STAT-5 (tyr(694)) was observed at 12h. Pharmacological inhibitors of p38 MAPK (SB203580) and JAK/STAT (AG490) but not ERK (PD98059) abrogated proliferation. RBL-induced expression of IL-2Rα and secretion of cytokines were drastically inhibited by SB203580 and AG490. CONCLUSIONS: RBL-induced proliferation and production of Th1/Th2 cytokines are mediated via p38 MAPK and STAT-5 signaling. GENERAL SIGNIFICANCE: RBL, a lectin with complex sugar specificity, is strongly mitogenic to human PBMC and stimulates the production of Th1 and Th2 cytokines. The results identified the signaling mechanism underlying the immunostimulatory activity of RBL.