Highly Purified Research Articles

Repository corticotropin injection (H.P. Acthar gel) for the treatment of sarcoidosis-induced hypercalciuria and vitamin D dysregulation: a pilot, open label study.

Background: Vitamin D dysregulation may occur in sarcoidosis patients and result in hypercalciuria, hypercalcemia, nephrolithiasis, and renal impairment. We performed an open label pilot study of highly purified (H.P.) Acthar® Gel (repository corticotropin injection) (RCI) on patients with sarcoidosis-induced vitamin D dysregulation and hypercalciuria. Methods: Nine patients with sarcoidosis-induced vitamin D dysregulation and hypercalciuria on stable maintenance anti-sarcoidosis therapy received 80 units of RCI subcutaneously twice weekly for 12 weeks. 24-hour urinary calcium excretion was measured at baseline and at 12 weeks. Other parameters measured over 16 weeks (including 4 weeks post the last RCI dose) included the following serum values: calcium, 25-OH vitamin D, 1,25- diOH vitamin D and serum parathyroid hormone (PTH). In addition, the Sarcoidosis Health Questionnaire (SHQ) and Short Form-36 (SF-36) as well as a urinary symptom score were measured in all subjects. Results: There was no significant change in the 24-hour urinary calcium excretion over 12 weeks of the study. However, there was evidence that RCI improved sarcoidosis-induced vitamin D dysregulation in that the serum 1,25- diOH vitamin D level significantly declined over 12 weeks. There was also improvement in most of the domains of the quality of life measures, although only a few of them reached statistical significance. There was also a trend toward improvement in urinary symptoms over the study period. The was evidence of the development corticosteroid side effects in the cohort, in that weight significantly increased over the study period. Conclusions: In this small pilot open label trial, 12 weeks of RCI did not significantly improve sarcoidosis-induced hypercalciuria. However, some statistically significant changes in serum vitamin D and PTH levels were demonstrated that were consistent with some amelioration of sarcoidosis-induced vitamin D dysregulation. Several corticosteroid-related side effects were demonstrated in this cohort. (Sarcoidosis Vasc Diffuse Lung Dis 2018; 35: 192-197).

Highly purified hMG versus recombinant FSH plus recombinant LH in intrauterine insemination cycles in women ≥35 years: a RCT.

Is the treatment with recombinant FSH (rFSH) plus recombinant LH (rLH) more effective than highly purified (HP)-hMG in terms of ongoing pregnancy rate (PR) in women ≥35 years of age undergoing intrauterine insemination (IUI) cycles? The ongoing PR was not significantly different in women treated with rFSH plus rLH or with HP-hMG. Although previous studies have shown beneficial effects of the addition of LH activity to FSH, in terms of PR in patients aged over 34 years having ovulation induction, no studies have compared two different gonadotrophin preparations containing LH activity in women ≥35 years of age in IUI cycles. A single-centre RCT was performed between May 2012 and September 2013 with 579 women ≥35 years of age undergoing IUI cycles. The patients were randomly assigned to one of the two groups, rFSH in combination with rLH group or HP-hMG (Meropur) group, by giving them a code number from a computer generated randomization list, in order of enrolment. The randomization visit took place on the first day of ovarian stimulation. Five hundred and seventy-nine patients with unexplained infertility or mild male factor undergoing IUI cycles were recruited in a university hospital setting. All women were enrolled in this study only for one cycle of treatment. Five hundred and seventy-nine cycles were included in the final analysis. Two hundred and ninety patients were treated with rFSH in combination with rLH and 289 patients were treated with HP-hMG. The ovarian stimulation cycle started on the third day of the menstrual cycle and the starting gonadotrophin doses used were 150 IU/day of rFSH plus 150 IU/day of rLH or 150 IU/day of HP-hMG. The drug dose was adjusted according to the individual follicular response. A single IUI per cycle was performed 34-36 h after hCG injection. The main outcome measures were ongoing PR and number of interrupted cycles for high risk of ovarian hyperstimulation syndrome (OHSS). Ongoing pregnancy rates were 48/290 (17.3%) in the recombinant group versus 35/289 (12.2%) in the HP-hMG group [(odds ratio (OR) 1.50, 95% CI 0.94-2.41, P = 0.09]. The number of interrupted cycles for high risk of OHSS was 13/290 (4.5%) in the rFSH plus rLH group and 2/289 (0.7%) in the HP-hMG group (OR 6.73, 95% CI 1.51-30.12, P = 0.013). One of the limitations of this study was the early closure and the ongoing PR could be overestimated. Both patient and gynaecologist were informed of the assigned treatment. Our results demonstrated the lack of differences in terms of ongoing PR between recombinant product and HP-hMG, in women ≥35 years undergoing controlled ovarian stimulation for IUI cycles. HP-hMG was safer than recombinant gonadotrophin concerning the risk of OHSS. None. NCT01604044.

Outcome from consecutive ICSI cycles in patients treated with recombinant human LH and those supplemented with urinary hCG-based LH activity during controlled ovarian stimulation in the long GnRH-agonist protocol

Clinical results were compared in a well-established, assisted reproduction program during the cross-over from highly purified (HP)-human menopausal gonadotropin (hMG) to rhFSH/rhLH. We included the last 33 patients treated with HP-hMG and the first 33 patients receiving rhFSH/rhLH for ovarian stimulation in their first intracytoplasmic sperm injection cycle. Patient baseline characteristics were almost identical in the two groups. Ovarian stimulation characteristics (days of stimulation, total amount of FSH administered using a modest initial loading dose of 150 IU/d, patients with oocyte retrieval) were similar for the two groups. However, the number of total and leading follicles and E2 serum levels on the human chorionic gonadotropin injection day were significantly higher in the rhFSH/rhLH group. The oocyte yield was significantly higher in the rhFSH/rhLH group as well as the number of metaphase II oocytes, difference almost reaching the statistical significance. The number of oocytes fertilized was also higher in patients receiving rhFSH/rhLH treatment. Implantation and clinical pregnancy rates were similar in both the study groups. It is concluded that in women undergoing controlled ovarian hyperstimulation under pituitary suppression for ART, the recombinant combined product containing FSH and LH in a fixed 2:1 ratio is more effective than HP-hMG in terms of follicle development, oocyte yield and quality, and fertilization rates.

IVF Outcomes with Either Highly Purified FSH vs. Recombinant FSH in Down-Regulated Normogonadotrophic Women: A Prospective Comparative Study in a Developing Country and Meta-Analysis

Objective: In order to compare the use of highly purified (HP) follicle stimulating hormone (FSH) with recom- binant FSH (recFSH) in a developing country, research should not only focus on clinical data, but also on the evidence available in the literature from previous trials. Study Design: We performed a prospective clinical trial with 118 infertile females undergoing IVF: HP-FSH (n = 59); recFSH (n = 59). In addition, we performed a meta-analysis of RCTs comparing currently available HP-FSH vs. recFSH. The primary outcomes for both studies were live-birth rate and rate of ovarian hyperstimulation. Results: In the clinical trial, the response to ovarian hyperstimulation was similar in both groups including the number of oocytes retrieved and the number of ampoules of gonadotrophins. The live birth rate per woman was 30.51% vs. 35.59% in the HP-FSH and recFSH treated groups respectively (P = 0.70). The rate of OHSS was 5.09% in the HP-FSH compared to 6.78% in the recFSH treated groups (P = 1.00). Regarding the meta-analysis, the live-birth (O.R= 1.30, 95% CI= 0.92 to 1.84) and OHSS rates (O.R= 1.14, 95% CI= 0.32 to 4.04) were not significantly different between the two groups. There was significantly less treatment days and total dose (IU) in the HP-FSH group compared with the rFSH group. Conclusion: HP-FSH yields similar clinical outcome to recFSH in terms of oocytes retrieved and live-birth rate.

A differential cytokine expression profile is induced by highly purified human menopausal gonadotropin and recombinant follicle-stimulating hormone in a pre- and postovulatory mouse follicle culture model

To compare the differential effects of highly purified (HP) hMG or recombinant FSH (rFSH) on cytokine expression before and after ovulation in an in vitro mouse ovarian follicle model. A prospective laboratory in vitro study. A university-based reproductive biology laboratory. MATERIAL(S): Mechanically isolated mouse preantral follicles from 14-day-old prepubertal mouse ovaries (F1 hybrids: C57BL/6JxCBA/ca). Randomly distributed mouse early preantral follicles were exposed to two hyperstimulation conditions with either HP-hMG or rFSH. An ovulatory stimulus was given using hCG/epidermal growth factor. Conditioned media from the two culture conditions were collected on the days before and after in vitro ovulation. Conditioned media were compared for their relative cytokine profile content as measured by a cytokine antibody array analysis. Relative concentrations of 62 cytokines in conditioned media before and after ovulation. Statistically significant increase in the production of a number of cytokines was found after HP-hMG stimulation compared with rFSH: 14 and 24 pre- and post-rhCG, respectively. Cytokines with the largest significant difference (more than 5 times) before and after ovulation included thymus-expressed cytokine (TECK), sTNFRI, and SDF-1alpha. The cytokines that are most strongly related to oocyte and embryo quality and implantation and that have been related to oocyte yield and maturation were significantly higher with HP-hMG. The significant differences in follicular cytokine production induced by HP-HMG and rFSH before and after in vitro ovulation might explain the difference in treatment outcome.

Highly purified hMG achieves better pregnancy rates in IVF cycles but not ICSI cycles compared with recombinant FSH: a meta-analysis

Objective. Human menopausal gonadotropin (hMG) was demonstrated to be superior to recombinant FSH (rFSH) regarding clinical outcomes. It is not clear whether this change in the evidence was due to the introduction of highly purified (HP) hMG.Design. Systematic review of properly randomised trials comparing HP-hMG vs. rFSH in women undergoing in vitro fertilisation (IVF) and/or intracytoplasmic sperm injection (ICSI). A meticulous search was performed using electronic databases and hand searches of the literature.Results. Six trials (2371 participants) were included. Pooling of the trials demonstrated that the probability of clinical pregnancy following HP-hMG administration was higher than rFSH and reached borderline significance (odd ratio (O.R) = 1.21, 95% confidence interval (CI) = 1.00 to 1.45), but the ongoing pregnancy/live-birth rate was not statistically different between the two drugs, although it showed strong trends towards improvement with HP-hMG (O.R = 1.19, 95% CI = 0.98 to 1.44).Subgroup analysis comparing both drugs in IVF cycles demonstrated a statistically significant better ongoing pregnancy/live-birth rate in favour of HP-hMG (O.R = 1.31, 95% CI = 1.02 to 1.68). On the other hand, there was almost an equal ongoing pregnancy/live-birth rate in ICSI cycles (OR = 0.98, 95% CI = 0.7 to 1.36).Conclusions. HP-hMG should be preferred over rFSH in women undergoing assisted reproduction, especially if IVF is the intended method of fertilisation.

Immunogenicity and efficacy of highly purified invasin complex vaccine from Shigella flexneri 2a

Development of a subunit vaccine for shigellosis requires identification of protective antigens and delivering these antigens in a manner that stimulates immunity comparable to that induced by natural infection. The Shigella invasin complex (Invaplex) vaccine is an ion-exchange-purified extract from virulent Shigella that consists of LPS and several other proteins, including the invasins IpaB and IpaC. Intranasal delivery of Invaplex stimulates protective immunity in small animal models for shigellosis. To identify the active component(s) of Invaplex responsible for its immunogenicity and efficacy, size-exclusion chromatography (SEC) was used to separate Invaplex into several different fractions. A high-molecular mass complex with a molecular mass between 669 MDa and 2 MDa consisted primarily of LPS, IpaB and IpaC and was considered to be a highly purified (HP) form of Invaplex. Using the mouse lung model to evaluate the immunogenicity and efficacy of the SEC fractions it was clearly demonstrated that the high-molecular mass complex of the invasins and LPS was responsible for the protective capacity of parent native Invaplex. Other smaller mass SEC fractions were mostly non-immunogenic and did not stimulate solid protection. In guinea pigs, the HP Invaplex stimulated an enhanced immune response as compared to the parent Invaplex and was fully protective. Isolation and characterization of the immunogenic and protective moiety within Invaplex will allow better standardization of the Invaplex product and may allow future development of an Invaplex assembled from purified components.

Influence of the cumulus cells exposed to exogenous luteinizing hormone (LH) activity during ovarian stimulation on human embryo development and intracytoplasmic sperm injection (ICSI) outcomes

Influence of the cumulus cells exposed to exogenous luteinizing hormone (LH) activity during ovarian stimulation on human embryo development and intracytoplasmic sperm injection (ICSI) outcomes

Efficacy and safety of highly purified hMG vs. recombinant follicle stimulating hormone: a meta-analysis

LH activity has been proposed to influence treatment response and outcome in assisted reproductive treatment. In order to assess its clinical profile and efficacy, we compared highly purified (HP) human menopausal gonadotrophin (hMG) (Menopur) with recombinant (r) FSH in IVF/ICSI cycles. Systematic review and meta-analysis of randomized trials. Meticulous computerized searches (last performed April 2007) were conducted by using MEDLINE, EMBASE, CENTRAL, LILACS, NRR, and trial registers of controlled trials. Furthermore, the reference lists of all known primary studies, review articles, citation lists of relevant publications, and included studies were examined to identify additional relevant citations. Finally, ongoing and unpublished trials were sought by contacting experts in the field and commercial entities. Primary outcome measures were the ongoing pregnancy/live birth rate and the rate of ovarian hyperstimulation syndrome. Secondary outcomes were the clinical pregnancy, multiple pregnancy and miscarriage rates, in addition to cycle characteristics (e.g. treatment duration, number of ampoules, E2 on day of HCG, number of oocytes retrieved, number of embryos obtained). The primary outcomes, ongoing pregnancy/live-birth (O.R = 1.19, 95% CI = 0.97 to 1.44) and OHSS rates (O.R = 1.19, 95% CI = 0.76 to 1.86) were not significantly different between the two groups. As for the secondary outcomes, there was significantly less treatment days (WMD = −1.21, 95% CI = −1.35 to −1.06), total dose (IU) (WMD = −285.61, 95% CI = −320.62 to −250.59), and embryos produced (WMD = −1.10, 95% CI = −1.80 to −0.40) in the rFSH group compared with the HP-hMG group. Even so, the number of oocytes retrieved (WMD = 0.83, 95% CI = 0.59 to 1.08) was significantly higher for the HP-hMG group. In addition, there was borderline significance with regards to the clinical pregnancy rate (O.R = 1.26, 95% CI = 1.00 to 1.58) also in favor of the HP-hMG group. The other secondary outcomes were not significantly different between the two groups. HP-hMG (Menopur) has been demonstrated to be as effacious and effective as rFSH with regards the clinical outcomes and patient safety.

Efficacy and safety of commercially available highly purified FSH vs. recombinant follicle stimulating hormone: a meta-analysis

Previous systematic reviews comparing highly purified (HP) follicle-stimulating hormone (FSH) with recombinant (r) FSH in IVF/ICSI cycles have demonstrated conflicting results. This may be related to different patient populations, study quality and individual drugs compared. Meta-analysis of randomized trials comparing only currently available HP-FSH vs. rFSH. In order to assess the clinical profile and efficacy of these drugs meticulous computerized searches (last performed April 2007) were conducted. Furthermore, the reference lists of all known primary studies, review articles, citation lists of relevant publications, and included studies were examined to identify additional relevant citations. Finally, ongoing and unpublished trials were sought by contacting experts in the field and commercial entities. Primary outcome measures were the ongoing pregnancy/live birth rate and the rate of ovarian hyperstimulation syndrome. Secondary outcomes were the clinical pregnancy, multiple pregnancy and miscarriage rates, in addition to cycle characteristics (e.g. treatment duration, number of ampoules, E2 on day of HCG, number of oocytes retrieved). The primary outcomes, ongoing pregnancy/live-birth (O.R = 1.24, 95% CI = 0.76 to 2.01) and OHSS rates (O.R = 6.70, 95%CI = 0.4 to 7.17) were not significantly different between the two groups. As for the secondary outcomes, there was significantly less treatment days (WMD = −0.43, 95% CI = −0.72 to −0.14) and total dose of FSH (IU) (WMD = −1172.29, 95% CI = −1445.13 to −899.45) in the HP-FSH group compared with the rFSH group. Even so, the number of oocytes retrieved (WMD = −0.46, 95% CI = −1.05 to 0.13) was not significantly different between the two groups. In addition, there were no significant differences with regards the clinical pregnancy rate (O.R = 1.34, 95% CI = 0.96 to 1.88) or the other secondary outcomes. HP-FSH has been demonstrated to be non-inferior to rFSH with regards the clinical outcomes and patient safety. In addition it was shown to be superior in stimulation efficacy than rFSH, with less numbers of ampoules and treatment days needed to attain similar results.

Bioactivity of a Highly Purified (HP) Urinary Follicle Stimulating Hormone (FSH) Evaluated by a Mouse Follicle Bioassay

FSH molecules show biochemical heterogeneity, reflected in the occurrence of isoforms exhibiting differences in physico-chemical and pharmacological properties. Dependent on their source, gonadotrophin preparations used in assisted reproductive technologies can differ regarding their clinical efficacy. Gonadotrophins target the ovary supporting and regulating follicle development, and determine steroid hormone production and oocyte maturation. In this study the bioactivity of a HP urinary FSH was assessed in comparison with a recombinant FSH preparation. The mouse follicle bioassay, a highly standardized in vitro model mimicking ovarian function, was used to investigate the impact of the gonadotrophins on folliculogenesis, steroidogenesis and oogenesis in a dose range covering sub- to supra-physiological conditions. Preantral ovarian follicles from pre-pubertal mice were individually cultured in 96-well plates during 12 days up to the pre-ovulatory stage. Meiosis resumed upon hCG stimulation, resulting in mature metaphase II oocytes on day 13. Follicles were exposed continuously to 1, 5, 10 and 100 IU/L of HP urinary FSH (Fostimon® = F) or recombinant FSH (Gonal F® = G). Per dose 60 follicles were cultured in 6 test replicates in 3 independent repeats. Follicle development was scored morphologically (follicle survival, differentiation stage, mucification of the cumulus-oocyte complex) and the steroid hormone profile was monitored in the medium throughout culture. Oocyte yield and nuclear maturation were analysed upon in vitro ovulation. Endpoint parameters are presented as mean values ± SD (P<0.05 was considered as significantly different). Follicle development, oocyte yield and maturation were similar for Fostimon® and Gonal F®. Folliculogenesis and oogenesis were not supported by the 1 IU/L dose, as follicles did not reach the antral stage and few oocytes extruded a polar body. When exposed to 5 IU/L FSH, 98±4% (F) and 92±10% (G) of the follicles survived, 83±16% (F) and 67±14% (G) developed an antral cavity and 95±8% (F) and 91±10% (G) responded to the HCG stimulus, whereas oocyte maturation rates were 94±6% (F) and 84±25% (G). Optimal follicle development was observed at 10 and 100 IU/L dose levels: > 90% of the follicles survived and formed an antrum; > 90% of the cumulus-oocyte complexes mucified in response to ovulatory stimulation; and ≥ 90% of the oocytes extruded a polar body. The hormone secretion profile was not significantly different for both preparations. Testosterone was measurable in all the replicates at the 100 IU/L dose level. Estradiol production increased dose-dependently, while progesterone secretion remained at basal levels throughout the follicular phase for 5 and 10 IU/L dose levels but increased in function of time at the 100 IU/L dose level. We demonstrated that equal doses of HP urinary Fostimon® and recombinant Gonal F® induced similar responses in the follicle bioassay. Folliculogenesis, oogenesis and steroidogenesis characteristics were comparable. The quantitative, multi-parametric test approach seemed highly effective to investigate gonadotrophin bioactivity at the target organ in a broad dose range, as the system allows the separate evaluation of gonadotrophin effects on different physiological processes in the ovary.

Economic evaluation of the administration of follitropin-β with a pen device

Previous studies suggest that administration of follitropin-β with a pen device (Puregon Pen®) is more convenient, less painful and 16–18% more efficient. The aim of this study was to perform an economic evaluation of the administration of follitropin-β by this pen device against follitropin-α by multidose and highly purified (HP) HMG by conventional syringe in IVF treatment by comparing the process utilities and the costs for the Dutch setting. Conjoint analysis assessed the process utilities for the three administration modes on a scale from 0 to 1. A decision analytic model estimated the costs of an average IVF cycle from a societal perspective. Patients estimated the process utility at 0.96 for the pen, 0.53 for the multidose and 0.36 for the conventional syringe. Additional costs were estimated at €0 and €194, comparing the pen with multidose or conventional methods respectively. Assuming a 16% efficiency gain of the pen, costs ranged from €–135 (savings) to €60 (extra costs). In conclusion, patients perceive sufficient benefits to the pen device to choose it over other dosing methods. Dominance of the pen device over the multidose method was shown. Compared with the conventional administration method, the added value of the pen device was 2.7 (0.96/0.36) times higher.

Highly purified (HP) Repronex® SC and HP-Repronex® IM are as effective as Repronex® SC in a prospective, randomized clinical trial evaluating controlled ovarian hyperstimulation (COH) in patients undergoing in-vitro fertilization (IVF)

Introduction: HMG has been successfully used in controlled ovarian hyperstimulation (COH) for nearly 40 years and remains the efficacy standard to which all other follicle stimulating agents are compared. Advanced manufacturing techniques have now produced the first ever, highly purified form of HMG (HP-Repronex; 75 IU FSH: 75 IU LH) that is nearly devoid of uncharacterized proteins.

Clinical efficacy of a highly purified SD‐treated factor IX concentrate prepared by conventional chromatography

Summary. A highly purified (HP) FIX concentrate was used in 12 patients with haemophilia B in order to prevent bleeding following surgery. The HP FIX concentrate (CRTS, Lille, France) was prepared by ion‐exchange chromatograpy and was SD treated. The patients underwent orthopaedic surgery (3), neurosurgery (3) or multiple dental extraction (6). They were treated for 2–18 days and received 8500 to 102000 FIX units. The mean FIX:C recovery was 0·92 ± 0·20%/U/kg. Satisfactory haemostasis was provided in all patients. None of them experienced thrombotic manifestations. There was no change in FDP or soluble fibrin complexes. Prothrombin fragments one plus two (F1 + 2), thrombin‐antithrombin III complexes (TAT) and cross‐linked fibrin degradation products (D‐dimer) remained in the normal range in one patient treated for 2·5 days for dental surgery. In contrast, elevated levels of F1+2, TAT and D‐dimer were observed in one patient undergoing a bilateral hip arthroplasty treated for 16 consecutive days with the same lot of HP FIX concentrate as the previous patient. There was no seroconversion for HIV, HBV and HCV. These results indicate that the HP FIX concentrate provides satisfactory haemostasis in surgery while avoiding excessive thrombogenic risks. However, some clinical circumstances, such as orthopaedic surgery, are known to be associated with especially high thrombotic risks that must be taken into account in case of FIX replacement.

Properties of a Highly Purified Human Plasma Factor IX:c Therapeutic Concentrate Prepared by Conventional Chromatography

We have characterized a highly purified (HP) factor IX concentrate intended for therapy of hemophilia B. The product has been prepared from pooled human plasma using a large-scale procedure combining three conventional chromatographic steps based on DEAE ion exchange and affinity on immobilized heparin. The specific activity of the product was 119±10 IU factor IX:c/mg protein (n = 15), corresponding to a purification factor of about 9,000. The concentrate was free of the vitamin K-dependent clotting factors II, VII and X and of proteins C and S. Most of the contaminants found in factor IX complex concentrate (PCC) were absent in this new product. High-molecular-weight kininogen, factors VIII, XI, XII or prekallikrein were not detected. There were no activated factors, such as factors IXa, and Xa, no thrombin and no phospholipids. Only two contaminants could be detected: C4 and inter-alpha-trypsin inhibitor (about 0.8 and 1.2mg/1,000IU factor IX:c, respectively). The purity of the product, as compared to PCC, was confirmed by sodium dodecylsulfate polyacrylamide gel electrophoresis, cellulose acetate electrophoresis, Grabar- Williams immunoelectrophoresis, and bidimensional immunoelectrophoresis. Thrombogenicity tests in rabbits revealed that the HP factor IX tested had a lower thrombogenic power than the PCC tested. The concentrate has been subjected to a 0.3% tri(n-butyl) phosphate-1% Tween 80 treatment for 6h at 25°C during its production to reduce or eliminate the risk of transmission of plasma-borne lipid-enveloped viruses. These conditions inactivated more than 3.8 log(10) of vesicular stomatitis virus and more than 4.3 log(10) of sindbis virus within 1 and 2 h of treatment, respectively. These data demonstrate that a highly purified therapeutic clotting factor IX concentrate can be prepared from human plasma by conventional chromatographic methods.

Relative cloning efficiencies and long-term propagation capacity for T cell clones of highly purified natural interleukin 2 compared to recombinant interleukin 2 in man

Highly purified (HP) natural (n) and recombinant (r) interleukin 2 (IL 2) have beencompared for their ability to support clonal outgrowth and long-term clonal propagation of alloactivated human T lymphocytes. The frequency of outgrowth of T cell colonies was as high as 1:2 to 1:4 when HPnIL 2 was employed for limiting dilution cloning. Separated CD4 + and CD8 + activated populations produced similar cloning efficiencies. Recloning of established CD4 + lines in HPnIL 2 suggested that a clonable cell frequency of 1:2–1:4 was the maximum possible that could be achieved in this system. In contrast, purified rIL 2 allowed outgrowth of only 1:10–1:20 cells from alloactivated populations. Again, CD4 + and CD8 + fractions generated similar cloning efficiencies. In terms of the fraction of derived clones, which could be propagated to > 1 × 10 6 and > 1 × 10 8 cells, nIL 2 again proved superior to rIL 2. In either source of IL 2, the proportion of CD4 + clones which could be extensively propagated was greater than in the CD8 + population. Surprisingly, although the addition of PHA to these lectin-free IL 2 preparations reduced the frequency of clonable cells, the proportion of clones which could be extensively propagated was increased. These results suggest that rIL 2, consisting of three differently glycosylated molecular species, may be preferable to rIL 2, which consists only of a single non-glycosylated species, for the cloning and long-term propagation of human T cells. These results may have some bearing on the choice of rIL 2 versus nIL 2 for therapeutic applications.

Interferon induces peripheral lymphadenopathy in mice.

Inoculation (i.p. or i.v.) of partially purified or highly purified (H.P.) mouse interferon (electrophoretically pure in SDS-PAGE gels) into different strains of mice (C3H, BALB/c, Swiss, heterozygote and athymic nu/nu C57BL/6) results in a marked increase in the weight and in the total number of cells in axillary and inguinal lymph nodes. Interferon injected into the hind footpad induced an increase in the weight of the ipsilateral popliteal lymph node compared with the contralateral lymph node. Inoculation of noninterferon proteins (impurities) obtained in the course of purification of this H.P. interferon, mock interferon, or human leukocyte interferon were without effect. Experiments were undertaken to elucidate the mechanism by which interferon induced an increase in the weight (number of cells) of peripheral lymph nodes. There was no evidence that the afflux of transferred chromium-labeled lymphoid cells to lymphoid tissue was increased in interferon-treated recipient mice. There was no evidence that interferon treatment resulted in a stimulation of the multiplication of cells within the peripheral lymph nodes. Since we observed a decrease in the number of lymphocytes in the thoracic duct lymph and in the peripheal blood of interferon-treated mice, we suggest that interferon treatment resulted in an inhibition of egress of lymphocytes from lymphoid tissue. The finding that interferon treatment of mice alters patterns of lymphocyte circulation may be relevant to some of the effects of interferon on the immune system and may explain in part the lymphadenopathy observed in the course of some viral infections.