Comparative sequence analysis in the exon 5 of growth hormone gene in the various livestock species of India.

The aim of the study was to identify genetic polymorphism in growth hormone (GH) gene locus of six different livestock species using PCR-Direct DNA sequencing method. In exon 5 of GH gene, 10 SNPs variants were identified in all livestock species studied, namely Bubalus bubalis, Bos indicus, Bos frontalis, Bos grunniens, Ovis aries, and Capra hircus. Four SNPs were observed in Bubalus bubalis, two SNPs in Bos indicus, one SNP in Ovis aries, and three SNPs in Capra hircus. No changes were observed in Bos grunniens and Bos frontalis when compared with the template sequence and the SNPs observed in the present investigation may be useful in the marker assisted selection.

Glyceraldehyde-3-phosphate dehydrogenase of the parasitic nematode Haemonchus contortus binds to complement C3 and inhibits its activity.

Haemonchus contortus is an economically important gastrointestinal parasite that infects primarily sheep and goats. To survive inside the host, the parasite must overcome the host immune response. In this study, we have identified and characterized a complement-C3-binding protein (H.c-C3BP) from this parasite employing biochemical and molecular biology tools. Initially, a truncated form of the protein was isolated from the excretory-secretory products of the parasite using C3-Sepharose column that facilitated its identification by mass spectroscopy. Subsequently, the parent molecule was generated in E. coli, and sequence analysis confirmed it as glyceraldehyde-3-phosphate dehydrogenase (GAPDH). GAPDH reacted with the antiserum raised against the truncated protein, and the truncated protein reacted with anti-GAPDH antiserum. The protein inhibited complement function as measured by haemolytic assay and membrane attack complex (MAC) formation. Sera from H. contortus-infected animals reacted with GAPDH as well as the truncated form of the protein, which further lend support to protein secretion. Thus, the C3-binding property of H. contortus GAPDH is a new function, and it represents a new entity of complement-binding protein. Identification and characterization of H.c-C3BP should facilitate development of new therapeutics considering a key role of this protein in immune modulation.

Evaluation of the effectiveness of diagnostic & management decision by teleophthalmology using indigenous equipment in comparison with in-clinic assessment of patients.

BACKGROUND & OBJECTIVES: There is a concern on the quality and the usefulness of teleophthalmology images, particularly those using indigenous equipment, in making a diagnosis and treatment decisions in ophthalmology. The present study was done to compare the level of agreement and sensitivity and specificity of diagnosis and management decisions of various eye diseases by teleophthalmology using indigenous equipment, compared to the in-clinic assessment. METHODS: Patients having different eye diseases were evaluated by two ophthalmologists - one ophthalmologist examined the patient in clinic setting while the other ophthalmologist made the diagnosis and management decision based on images sent by teleophthalmology. The images were taken by the ophthalmic technician using digital imaging system and fundus camera. The clinical findings and management decisions by the two ophthalmologists were masked to each others. RESULTS: In diagnosis of anterior segment eye diseases such as cataract and corneal diseases there was good to very good agreement (kappa values of 0.68 and 0.91 for cataract and corneal diseases respectively) between in-clinic assessment and assessment by teleophthalmology. There was moderate agreement (kappa values of 0.52 and 0.48 for glaucoma and retinal diseases respectively) between in-clinic assessment and assessment by teleophthalmology for the diagnosis of glaucoma and retinal diseases. For the management decisions of patients, there was moderate level of agreement in all groups of eye diseases. INTERPRETATION & CONCLUSIONS: Teleophthalmology, using indigenous equipment was found to be effective in diagnosis and management decision of anterior segment eye diseases such as cataract and cornea, and with some modification and continuous training to the technicians could become an effective tool for screening and referral of glaucoma and retinal diseases.

Molecular characterization and serodiagnostic evaluation of the Echinococcus ortleppi recombinant glutaredoxin 1 protein for cystic echinococcosis in buffalo (Bubalus bubalis).

Cystic echinococcosis (CE), caused by the metacestode of Echinococcus granulosus sensu lato (s.l.), adversely affects the physiology of the vital organs in which they grow. Condemnation of meat causes substantial economic loss to the livestock industry. Conventionally the infection is detected by necropsy as serological diagnosis of the infection in livestock is ambiguous. Identification of specific diagnostic antigens would be a substitute for the cyst fluid antigens which lack adequate diagnostic sensitivity and specificity. BLAST analysis supported by the negligible pairwise nucleotide distance of the 389 nt COX1, 489 nt NAD1, and 425 nt ITS1 with the related sequences of E. ortleppi ascertained the association of E. ortleppi with CE in buffaloes. Given the extensive distribution of glutaredoxin 1 in every developmental stage of Echinococcus granulosus s.l that makes it an ideal serodiagnostic antigen for CE, we expressed the 14 kDa E. ortleppi glutaredoxin 1 (rEoGrx1) protein in E. coli BL21 (DE3) and tested a total of 225 sera samples, including 126 sera samples from the necropsy-positive buffalo, by the rEoGrx1 IgG-ELISA. The ELISA could detect a total of 82/126 sera samples as positive. The diagnostic sensitivity and specificity of the rEoGrx1 IgG-ELISA were 65.1 % and 51.5 %, respectively. The protein showed serological cross-reaction against Fasciola gigantica, Toxoplasma gondii, and Sarcocystis sp. The in silico bioinformatics analysis of the E. ortleppi, F. gigantica, and T. gondii glutaredoxin sequences revealed fully conserved amino acids at positions 11 and 21, the substitution of conserved amino acids at positions 14 and 6, and semi-conserved substitutions at positions 3 and 4, respectively. The findings partly explain the molecular basis of the serological cross-reactivity of the protein.

Vaccination of buffaloes with Fasciola gigantica recombinant glutathione S-transferase and fatty acid binding protein.

Fasciola gigantica, causative agent of tropical fasciolosis, inflicts substantial economic losses on the livestock industry, affecting severely buffalo productivity in the tropical countries. Very few vaccination trials with different target antigens against F. gigantica infection have been conducted in this host. Present study describes a vaccination trial in buffaloes with F. gigantica recombinant glutathione S-transferase and fatty acid binding protein. The two recombinant proteins were expressed in Escherichia coli and evaluated for their immunoprophylactic potential in buffalo calves, using montanide 70 M-VG, a mineral oil-based adjuvant, for delivering the antigens. Buffalo calves were distributed in three groups, with group I, II and III calves immunized with recombinant glutathione S-transferase, fatty acid binding protein and a cocktail of these two antigens, respectively. Immunization of the calves evoked a mixed IgG1 and IgG2 antibody response. Present vaccination trial in these animals achieved a maximum protection level of 35%, when the two antigens were used in combination. Eosinophils were measured in both immunized and non-immunized challenge control animals, which showed a steady increase in their count in response to immunization with both the antigens and infection with F. gigantica, respectively.

Lidar quantification of bank erosion in Blue Earth County, Minnesota.

Sediment and phosphorus (P) transport from the Minnesota River Basin to Lake Pepin on the upper Mississippi River has garnered much attention in recent years. However, there is lack of data on the extent of sediment and P contributions from riverbanks vis-à-vis uplands and ravines. Using two light detection and ranging (lidar) data sets taken in 2005 and 2009, a study was undertaken to quantify sediment and associated P losses from riverbanks in Blue Earth County, Minnesota. Volume change in river valleys as a result of bank erosion amounted to 1.71 million m over 4 yr. Volume change closely followed the trend: the Blue Earth River > the Minnesota River at the county's northern edge > the Le Sueur River > the Maple River > the Watonwan River > the Big Cobb River > Perch Creek > Little Cobb River. Using fine sediment content (silt + clay) and bulk density of 37 bank samples representing three parent materials, we estimate bank erosion contributions of 48 to 79% of the measured total suspended solids at the mouth of the Blue Earth and the Le Sueur rivers. Corresponding soluble P and total P contributions ranged from 0.13 to 0.20% and 40 to 49%, respectively. Although tall banks (>3 m high) accounted for 33% of the total length and 63% of the total area, they accounted for 75% of the volume change in river valleys. We conclude that multitemporal lidar data sets are useful in estimating bank erosion and associated P contributions over large scales, and for riverbanks that are not readily accessible for conventional surveying equipment.

Chickpea (Cicer arietinum L.) as model legume for decoding the co-existence of Pseudomonas fluorescens and Mesorhizobium sp. as bio-fertilizer under diverse agro-climatic zones.

Microbial co-inoculation strategy utilizes a combination of microbes to stimulate plant growth concomitant with an increased phytopathogen tolerance. In the present study, 15 endophytic bacterial isolates from rhizosphere and roots of wild chickpea accessions (Cicer pinnatifidum, C. judiacum, C. bijugum and C. reticulatum) were characterized for morphological, biochemical and physiological traits. Two promising isolates were identified as Pseudomonas fluorescens strain LRE-2 (KR303708.1) and Pseudomonas argentinensis LPGPR-1 (JX239745.1) based on 16S rRNA gene sequencing. Biocompatibility of selected endophytes with Mesorhizobium sp. CH1233, a standard isolate used as a national check in All India Coordinated Research Project (AICRP) was assessed to develop functional combinations capable of producing Indole acetic acid, gibberellins, siderophores and improving seed vigour (in vitro). In vivo synergistic effect of promising combinations was further evaluated under national AICRP, (Chickpea) at two different agro-climatic zones [North-West plain (Ludhiana and Hisar) and Central zones (Sehore)] for three consecutive Rabi seasons (2015-18) to elucidate their effect on symbiotic, soil quality and yield parameters. On the pooled mean basis across locations over the years, combination of Mrh+LRE-2 significantly enhanced symbiotic, soil quality traits and grain yield over Mrh alone and highly positive correlation was obtained between the nodulation traits and grain yield. Superior B: C ratio (1.12) and additional income of Rs 6,505.18 ha-1 was obtained by application of Mrh+LRE-2 over Mrh alone and un-inoculated control. The results demonstrate that dual combination of Mrh and Pseudomonas sp. from wild Cicer relatives can be exploited as a potential bio-fertilizer for increasing soil fertility and improving chickpea productivity under sustainable agriculture.

Efficacy of hydrocortisone acetate/hyaluronidase vs triamcinolone acetonide/hyaluronidase in the treatment of oral submucous fibrosis.

BACKGROUND & OBJECTIVES: Oral submucous fibrosis is a common premalignant condition caused by chewing arecanut and other irritants in various forms. Its medical treatment is not yet fully standardized, although the optimal doses of its medical treatment is in the form of hydrocortisone acetate combined with hyaluronidase. The problem with the prevailing treatment was injections at weekly interval. In this study we compared the efficacy of hydrocortisone acetate and hyaluronidase at weekly interval versus triamcinolone acetonide and hyaluronidase at 15 days interval. METHODS: Patients of OSMF (100) were randomly divided into two groups A and B. Group A patients received combination of hydrocortisone acetate (1.5 ml)/hyaluronidase (1500 IU) at weekly interval submucosally in pterygomandibular raphe, half dose on each side for 22 wk. Group B patients received combination of triamcinolone acetonide (10 mg/ml)/ hyaluronidase (1500 IU) at 15 days interval for 22 wk. Treatment outcome was evaluated on the basis of improvement in symptom score, sign score and histopathological improvement. Student's 't' test was applied for comparing the results. RESULTS: No statistically significant difference in symptom score, sign score and histopathological improvement was seen between the two groups. INTERPRETATION & CONCLUSION: Treatment regimen of group B was more convenient to the patients because less number of visits required and cheap. No side effects were seen. A follow up study is required to see long term effects.

Effect of FSH and LH hormones on oocyte maturation of buffalo and gene expression analysis of their receptors and Cx43 in maturing oocytes.

Follicle stimulating hormone (FSH) and luteinizing hormone (LH) are commonly added to maturation media to improve cumulus expansion known to be a predictor of oocyte maturation. Therefore, effects of various concentrations of FSH (1000 ng/ml), LH (1000 ng/ml) and FSH + LH (1000 ng/ml each) in comparison with control (without FSH + LH) cultured oocytes were investigated. FSH and LH (1000 ng/ml each) induced significantly more cumulus expansion and polar body numbers, as compared with control and treatments of 1000 ng/ml FSH and 1000 ng/ml LH alone. Expression of FSH receptor (r), LHr and Cx43 mRNAs was determined by real-time PCR in cumulus-oocyte complexes (COCs) and denuded oocytes at different maturation times. Expression of all three genes was higher in COCs compared with denuded oocytes, confirming the importance of cumulus cells in oocyte maturation. FSHr and connexin 43 (Cx43) mRNA abundance in both COCs and denuded oocytes was highest at 0-6 h of maturation and decreased subsequently. However, LHr mRNA abundance increased from 6 h up to 24 h of maturation. The study concluded that FSH, LH receptors and Cx43 gene expression regulation is an index related to oocyte maturation.

Efficiency of SCNT buffalo (Bubalus bubalis) embryos in different culture medium and analysis of mRNA expression of insulin-like growth factors during embryogenesis.

Growth factors in culture media are known to affect the embryo production rates in in vitro production cultures. To improve the efficiency of somatic cell nuclear transfer (SCNT) derived embryos in Indian buffaloes (Bubalus bubalis), embryos were cultured in three different culture mediums viz. Group-A; TCM-199 + FBS, Group-B; TCM-199 + Poly vinyl alcohol (PVA) and Group-C; CR1aa + BSA. Embryo production rate and expression level of insulin-like growth factor genes (IGF-1, IGF-1R, IGF-2 and IGF-2R) were analysed in embryo culture. Cleavage and blastocyst production rates were 62.5% and 22.3% in Group-A, 53.8% and 13.0% in Group-B and 62.0% and 19.2% in Group-C respectively, whereas in in vitro fertilization (IVF) control cultured in TCM-199 plus 10% FBS, rates were 79.1% and 29.4%. Relative gene expression of SCNT embryos was compared with that in IVF control. IGF-1 and IGF-2 mRNA expression at blastocyst stage was up-regulated (p ≤ 0.05) in all culture groups, while IGF-1R and IGF-2R expression was down regulated (p ≤ 0.05) in Group-B and Group-C. In conclusion, the higher mRNA levels at certain stages in different culture conditions affected in vitro development of SCNT embryos. These results show that the transcript level of the insulin-like growth factor genes was significantly altered by in vitro culture condition. Culture medium TCM-199 with 10% FBS produced higher number of embryos and was able to co-op with gene expression of IVF control. Differences continue to be observed between SCNT cultured and IVF embryos, and until these differences are minimized, aberrations in SCNT embryonic development will continue to arise.

Molecular cloning and characterization of a glutathione S-transferase in the tropical liver fluke, Fasciola gigantica.

Glutathione S-transferase from an Indian isolate of Fasciola gigantica of buffalo origin was isolated and characterized. Total RNA was transcribed to cDNA by reverse transcription and an amplicon of 657 bp glutathione S-transferase gene was obtained by polymerase chain reaction (PCR). The present isolate showed 99.1% sequence homology with the published sequence of the F. gigantica GST gene of cattle origin, with six nucleotide changes causing an overall change of four amino acids. Glutathione S-transferase protein was expressed in Escherichia coli using a prokaryotic expression vector pPROEXHTb. The recombinant protein was purified under non-denaturing and denaturing conditions by nickel nitrilotriacetic acid (Ni-NTA) affinity chromatography. Recombinant GST protein detected F. gigantica infection in naturally infected buffaloes by dot-ELISA.

Bullet and Blast ENT Injuries in Counter-Insurgency Area.

BACKGROUND: Many ENT injuries are not recognized easily, but they have the potential of increasing the morbidity. METHODS: ENT injuries managed in two tertiary care Level-IV hospitals between 2006 and 2007 were studied with a view to formulate strategy in efficient management of these cases. RESULT: Emergency bags did not carry readymade packs to control nasal bleeds. Routinely screening of ears in all blast injury cases in the 'Blast Injury Program' helped in early identification of hearing loss. Lack of sufficient stenting of nasal cavities resulted in severe nasal stenosis which was difficult to repair. Splinters lodged in pharyngeal wall escaped detection, resulting in concealed haemorrhage and shock. CONCLUSION: Nasal packs and epistaxis catheters must be included in emergency bags to minimize blood loss at first contact. Screening for ear trauma in all blast injuries increases detection rate and is beneficial to the soldier. Stenting of injured nasal cavities and early transfer to a tertiary care hospital could reduce morbidity. Plain radiography of head and neck areas could help detect splinters in vital areas and guide management.

PLGA nanoparticles stabilized with cationic surfactant: safety studies and application in oral delivery of paclitaxel to treat chemical-induced breast cancer in rat.

PURPOSE: This study was carried out to formulate poly(lactide-co-glycolide) (PLGA) nanoparticles using a quaternary ammonium salt didodecyl dimethylammonium bromide (DMAB) and checking their utility to deliver paclitaxel by oral route. METHODS: Particles were prepared by emulsion solvent diffusion evaporation method. DMAB and particles stabilized with it were evaluated by MTT and LDH cytotoxicity assays. Paclitaxel was encapsulated in these nanoparticles and evaluated in a chemical carcinogenesis model in Sprague Dawley rats. RESULTS: MTT and LDH assays showed the surfactant to be safe to in vitro cell cultures at concentrations <33 microM. PLGA nanoparticles prepared using this stabilizer were also found to be non-toxic to cell lines for the duration of the study. When administered orally to rats bearing chemically induced breast cancer, nanoparticles were equally effective/better than intravenous paclitaxel in cremophor EL at 50% lower dose. CONCLUSIONS: This study proves the safety and utility of DMAB in stabilizing preformed polymers like PLGA resulting in nanoparticles. This preliminary data provides a proof of concept of enabling oral chemotherapy by efficacy enhancement for paclitaxel.

Relative expression of cell growth regulatory genes insulin-like growth factors (IGF-1 and IGF-2) and their receptors (IGF-1R and IGF-2R) in somatic cell nuclear transferred (SCNT) and in vitro fertilized (IVF) pre-implantation buffalo embryos.

Relative mRNA transcript expression of insulin-like growth factors, IGF-1, IGF-2 and their receptors, IGF-1R and IGF-2R, was studied in SCNT and IVF buffalo embryos at different developmental stages using SYBR green with real-time PCR. SCNT embryos were produced by enucleating IVM oocytes and transferring granulosa cells (passage 5) followed by the electrofusion and chemical activation method. IVF embryos were produced by culturing 15-20 COCs in BO capacitated sperms from frozen and thawed buffalo semen. SCNT embryo production rate was lower than IVF. IGF-1 mRNA expression was significantly upregulated at 2-cell, 16-cell, morula and blastocyst stages of SCNT embryos than in IVF embryos. IGF-1R expression declined from the 2-cell to the 16-cell stage, which increased in later stages and was highest in IVF blastocysts. Similar regulation was observed at different stages in SCNT embryos, except at the 4-cell stage where expression was higher. IGF-2 expression decreased up to the 8-cell stage and increased until the blastocyst stage, being higher in SCNT than IVF embryos. IGF-2R mRNA transcript expression was consistently lower in SCNT than in IVF embryos. Reprogramming of IGF-1, IGF-1R, IGF-2 and IGF-2R expression may have a significant role in cell proliferation in cloned embryos and is developmentally regulated.

Lipid profile in oral submucous fibrosis.

BACKGROUND: Changes in lipid profile have long been associated with malignancies as lipids play a key role in maintenance of cell integrity. This study evaluated the alterations in extended lipid profile in untreated patients of oral submucous fibrosis (OSMF) and studied the correlation between lipid levels with tobacco consumption. MATERIALS AND METHODS: In this hospital-based study, 65 clinically diagnosed and histopathologically proven patients of OSMF and 42 age and sex matched controls were studied. In these samples serum lipids including: (i) Total cholesterol, (ii) LDL cholesterol (LDLC), (iii) HDL cholesterol (HDLC) (iv) VLDL cholesterol (VLDLC) (v) triglycerides (vi) Apo-A1 (viii) Apo-B and (viii) LPa were analyzed. RESULTS: A significant decrease in plasma total cholesterol, HDLC and Apo-A1 was observed in patients with OSMF as compared to the controls. Thus an inverse relationship between plasma lipid levels and patients was found in OSMF. CONCLUSION: The lower levels of plasma cholesterol and other lipid constituents in patients might be due to their increased utilization. The findings strongly warrant an in-depth study of alterations in plasma lipid profile in patients with oral precancerous conditions.

Improvement of in vitro oocyte maturation with lectin supplementation and expression analysis of Cx43, GDF-9, FGF-4 and Fibronectin mRNA transcripts in Buffalo (Bubalus bubalis).

PURPOSE: To optimize In vitro maturation (IVM) of quality oocytes for embryo production through IVF and SCNT. METHODS: Buffalo oocytes were in vitro matured in the presence of the pokeweed lectin (Phytolacca americana), a potent lymphocyte mitogen. Lectin was supplemented in TCM + 10% FBS at the doses of 0, 1, 5, 10, 15, 20 and 40 microg/ml and cumulus expansion and gene expression patterns were characterized. RESULTS: The degree of cumulus expansion in different lectin treatment levels improved from 1.1 at 1 Ag/ml level to 3.60 at 10 microg/ml level and then decreased in higher concentration 20 microg/ml (1.66) and 40 microg/ml (0.64). IVF embryos showed highest cleavage rate (88.8%) in 10 microg/ml lectin treatment. Expression of all mRNA transcript studied (Cx43, GDF 9, FGF-4 and Fibronectin) was positively correlated with cumulus expansion and polar body extrusion. CONCLUSIONS: Mitogenic lectin supplemented maturation media improves oocyte quality for in vitro embryo production.

Laparoscopic cholecystectomy under spinal anesthesia: a study of 3492 patients.

OBJECTIVES: Spinal anesthesia (SA) for laparoscopic cholecystectomy (LC) is only contemplated in patients where general anesthesia (GA) is contraindicated. In this paper, we present our experience of over 12 years of performing laparoscopic cholecystectomy, primarily under spinal anesthesia. METHODS: Over the last 12 years, LC was done under SA in 3492 patients. There was no modifications in technique, as compared to the same procedure done under GA. The intra-abdominal pressure was kept at 8-10 mm Hg. Sedation was given, if required, and conversion to GA was done in patients not responding to sedation or with failure of SA. RESULTS were compared with those of 538 patients undergoing LC under GA. RESULTS: Eighteen (0.52%) patients required a conversion to GA. Hypotension requiring support was recorded in 700 (20.05%) patients, and 429 (12.29%) patients experienced neck and/or shoulder pain. Overall, 2.29% (80) patients had vomiting in the postoperative period, as compared to 30.30% (163 patients) in patients administered GA. In total, 34.36% (1200) patients required injectable diclofenac for their abdominal pain within 2 hours postoperatively after SA, and oral analgesic was required in 2150 (61.57%) patients within the first 24 hours. On the other hand, 91.45% patients operated on under GA required injectable analgesics in the immediate postoperative period. Postural headache persisting for an average of 2.6 days was seen in 206 (5.9%) patients in the postoperative period after SA. There was no difference in operative parameters between the two groups, with 88.95% patients requiring only three ports under SA, as compared to 89.41% in the GA group. Operative time, too, was not much different. Postoperatively, biliary leak was seen in 0.60% patients, as compared to 0.93% in the GA group. Average time to discharge was almost similar in both the groups. Kernofsky's performance status showed a 98.6% satisfaction level in patients. CONCLUSIONS: LC done under spinal anesthesia does not require any change in technique and, at the same time, has a number of advantages, as compared to general anesthesia, and should be the anesthesia of choice.

Immune responses to polyethylenimine delivered plasmid DNA encoding a Fasciola gigantica fatty acid binding protein in mice and rabbits.

Fasciola gigantica fatty acid binding protein (FABP) was evaluated for evoking an effective immune response in mice and rabbits, when delivered as a DNA vaccine in muscle cells. Polyethylenimine (PEI), 25 kDa, branched cationic polymer was used as a delivery vehicle for this DNA in the muscle cells of mice and rabbits. Naked DNA evoked mixed Th1 and Th2 responses in mice. PEI condensed DNA, at amine nitrogen over DNA phosphate (N/P) ratios of 4, 6 and 8 and with various DNA concentrations, failed to evoke a significantly higher antibody response compared to naked DNA in mice. Similarly, the humoral immune response to naked DNA administration in rabbit thigh muscles was poor and no boosting of this antibody response on administration of DNA complexed to PEI was observed. On metacercarial challenge, rabbits failed to show any significant protective immune response in both the naked DNA and PEI-DNA immunized groups. Administration of PEI alone (12.5 mug) in mouse thigh muscles caused significant muscle cytotoxicity but condensation of DNA with PEI had less of a toxic effect on muscle cells, which was inversely related to the N/P ratio. Delivery of plasmid DNA encoding F. gigantica FABP by high molecular weight polyethylenimine (branched, 25 kDa) did not boost the effective immune response in both the animal species, which could either be attributed to cytotoxicity associated with this cationic polymer or muscle cells being unsuitable target cells for PEI condensed DNA delivery.