Amjad Riaz

Artemisinins and immune system.

Artemisinins in combination with other antimalarial drugs remain the mainstay of current antimalarial armamentarium. It is interesting to note that many traditional drugs with antiprotozoal background can wield immunomodulation on the recipients immune system in a positive or negative direction. Artemisinins also attribute immunomodulatory distensions. For instance, they demonstrate predominant immunosuppressive traits toward different immune components by particularly regulating the cellular proliferation and cytokine release, which indicates that they possess some additional mechanisms and features demanding deliberate attentions. This article reviews the data-based immunomodulatory effects of artemisinins on different immune cells including neutrophils, macrophages, splenocytes, T and B cells in conjunction with their therapeutic prospective with regard to inflammation, autoimmunity and delayed-type hypersensitivity.

Mouse cloning and somatic cell reprogramming using electrofused blastomeres.

Mouse cloning from fertilized eggs can assist development of approaches for the production of “genetically tailored” human embryonic stem (ES) cell lines that are not constrained by the limitations of oocyte availability. However, to date only zygotes have been successfully used as recipients of nuclei from terminally differentiated somatic cell donors leading to ES cell lines. In fertility clinics, embryos of advanced embryonic stages are usually stored for future use, but their ability to support the derivation of ES cell lines via somatic nuclear transfer has not yet been proved. Here, we report that two-cell stage electrofused mouse embryos, arrested in mitosis, can support developmental reprogramming of nuclei from donor cells ranging from blastomeres to somatic cells. Live, full-term cloned pups from embryonic donors, as well as pluripotent ES cell lines from embryonic or somatic donors, were successfully generated from these reconstructed embryos. Advanced stage pre-implantation embryos were unable to develop normally to term after electrofusion and transfer of a somatic cell nucleus, indicating that discarded pre-implantation human embryos could be an important resource for research that minimizes the ethical concerns for human therapeutic cloning. Our approach provides an attractive and practical alternative to therapeutic cloning using donated oocytes for the generation of patient-specific human ES cell lines.

Effect of collection frequency on the semen quality of broiler breeder

1. Seven 35-week-old Hubbard broiler breeder males were subjected to three semen collection frequencies either once every 2 d (48 h), daily (24 h) or twice daily (12 h). 2. Semen characteristics including motility, volume, concentration and sperm numbers per ejaculate were determined for each ejaculate. 3. Sperm motility was unaffected by collection interval, but semen volume was lower at 12 than 24 h intervals. Sperm concentration was lower at 12 than 48 h intervals. 4. At 24 and 48 h number of sperm per collection (1·7 ± 0·2, 1·8 ± 0·2 × 109) were higher than at 12 h (1·2 ± 0·1 × 109). 5. The number of semen doses over a 6-d period increased linearly as the frequency of collection increased from once every 2 d to twice daily. 6. It is concluded that output was theoretically maximal at twice daily collection, but in practice not all cockerels may be able to maintain full performance with such a demanding regime.

Trehalose improves semen antioxidant enzymes activity, post-thaw quality, and fertility in Nili Ravi buffaloes (Bubalus bubalis).

Our objectives were to study the effect of trehalose in extender on (1) antioxidant enzymes profile during cryopreservation (after dilution, before freezing, and after thawing), (2) in vitro quality (after thawing), and (3) in vivo fertility of Nili Ravi buffalo (Bubalus bubalis) bull spermatozoa. Semen samples (n = 20) from four buffalo bulls were diluted in Tris-citric acid-based extender having different concentrations of trehalose (0.0, 15, 30, 45, and 60 mM) and frozen in French straws. At post dilution, profile of sperm catalase (U/mL) was higher (P < 0.05) in extenders containing 15, 30, and 45 mM of trehalose as compared to control. Although profiles of superoxide dismutase (U/mL) and total glutathione (μM) were higher (P < 0.05) in extenders containing 15 and 30 mM of trehalose as compared to control. At prefreezing, sperm catalase, superoxide dismutase, and total glutathione profiles were higher (P < 0.05) in all the treatment groups as compared to control. At post thawing, the profiles of catalase and total glutathione were higher (P < 0.05) in extender containing 30-mM trehalose as compared to other treatment groups and control. Whereas, profile of superoxide dismutase was higher (P < 0.05) in extenders containing 30, 45, and 60 mM of trehalose as compared to control and 15mM group. Post thaw total sperm motility (%) was higher (P < 0.05) in extender containing 30-mM trehalose as compared to control and 15 and 60-mM groups. Although sperm progressive motility (%), rapid velocity (%), average path velocity (μm/s), straight line velocity (μm/s), curvilinear velocity (μm/s), plasma membrane (structural and functional, %), acrosome (%), and DNA (%) integrity were higher (P < 0.05) in extender containing 30 mM trehalose as compared to other treatment groups and control. The fertility rates (61% vs. 43%) were higher (P < 0.05) in buffaloes inseminated with semen doses cryopreserved in extender containing 30 mM of trehalose than the control. It is concluded that addition of 30-mM trehalose in extender improves the semen antioxidant enzymes activity, post thaw quality, and fertility in Nili Ravi buffaloes.

Sperm survival kinetics in different types of bull semen: progressive motility, plasma membrane integrity, acrosomal status and reactive oxygen species generation

This study was designed to compare the kinetics of sperm survival in different types of bull semen. Fresh ejaculates from four bulls were pooled, diluted in Tris-citric acid-egg yolk-glycerol extender, cooled to 4°C, frozen in LN2 and thawed at 37°C. Fresh, diluted, cooled and frozen-thawed semen were incubated at 37°C, and evaluated at 0, 2, 4, 6, 12 and 24h after the beginning of incubation. In Experiment 1, progressive sperm motility, normal acrosomes and plasma membrane integrity and asymmetry were determined. In Experiment 2, generation of superoxide anion (O2(•)) along with plasma membrane permeability and generation of hydrogen peroxide (H2O2) along with plasma membrane integrity were assessed. In Experiment 1, frozen-thawed semen had shorter survival times for progressive sperm motility, and spermatozoa with intact plasma membranes and acrosomes (IPM-IACR) as compared with other types of semen (P<0.05). Fresh spermatozoa underwent a necrotic pathway, diluted and cooled spermatozoa underwent an apoptosis-like pathway and frozen-thawed spermatozoa underwent both necrotic and apoptosis-like pathways. In Experiment 2, spermatozoa in all four types of semen exhibited O2(•-) generation and increased plasma membrane permeability, and became necrotic without H2O2 generation during incubation (P<0.05). In conclusion, frozen-thawed semen had shorter sperm longevity, which has important implications relating to the timing of artificial insemination. Different types of semen followed different death pathways. During incubation, spermatozoa in all types of semen generated O2(•-), which increased the permeability and compromised the integrity of the plasma membrane.

l-Cysteine improves antioxidant enzyme activity, post-thaw quality and fertility of Nili-Ravi buffalo (Bubalus bubalis) bull spermatozoa

The effects of l‐cysteine in extender on antioxidant enzymes profile during cryopreservation, post‐thaw quality parameters and in vivo fertility of Nili‐Ravi buffalo bull spermatozoa were studied. Semen samples from 4 buffalo bulls were diluted in Tris–citric acid‐based extender having different concentrations of l‐cysteine (0.0, 0.5, 1.0, 2.0 and 3.0 mm) and frozen in 0.5‐ml French straws. The antioxidative enzymes [catalase, super oxide dismutase and total glutathione (peroxidase and reductase)] were significantly higher (P < 0.05) at pre‐freezing and post‐thawing in extender containing 2.0 mm l‐cysteine as compared to other groups. Post‐thaw total motility (%), progressive motility (%), rapid velocity (%), average path velocity (μm s−1), straight line velocity (μm s−1), curvilinear velocity (μm s−1), beat cross frequency (Hz), viable spermatozoa with intact plasmalemma (%), acrosome and DNA integrity (%) were higher with the addition of 2.0 mm l‐cysteine as compared to other groups (P < 0.05). The fertility rates (59 versus 43%) were higher (P < 0.05) in buffaloes inseminated with doses containing 2.0 mm of l‐cysteine than in the control. In conclusion, the addition of 2.0 mm l‐cysteine in extender improved the antioxidant enzymes profile, post‐thaw quality and in vivo fertility of Nili‐Ravi buffalo bull spermatozoa.

Polarity-Based Solvents Extraction of Opuntia dillenii and Zingiber officinale for In Vitro Antimicrobial Activities

Extracts from dried stem of Opuntia dillenii and rhizome of Zingiber officinale were evaluated for antimicrobial activities by extraction in non-polar (petroleum ether and chloroform) and polar solvents (methanol and water). Bacillus subtilis and Staphylococcus aureus showed considerable susceptibility to all extracts of Opuntia dillenii and Zingiber officinale. Ether and chloroform extracts of Opuntia dillenii showed improved antimicrobial activity against Escherichia coli (gram negative) as compared to that of its methanolic and water extracts. On the other hand, methanolic and water extracts of Zingiber officinale showed better susceptibility for Escherichia coli. Salmonella typhi (gram negative) was found to be resistant to all extracts of both Opuntia dillenii and Zingiber officinale. Our results indicated that both Opuntia dillenii and Zingiber officinale have potent antimicrobial activity that is also based on solvent polarity during extraction.

Aspirin may do wonders by the induction of immunological self-tolerance against autoimmune atherosclerosis

Induction of immune tolerance is one of the recent novel immunomodulatory strategies to directly intervene the autoimmune-driven atherosclerosis. Aspirin is a prototypic non-steroidal anti-inflammatory drug, which is now being regarded as a life-saver in variety of atherosclerotic cardiovascular complications. Considerable amount of data emerged during last few years clearly suggests that aspirin can cause immunomodulation by several mechanisms, particularly, its ability to induce tolerogenic dendritic cells (DCs) and to upregulate T regulatory (Treg) cells is especially appealing with respect to induction of immunological self-tolerance. Based on this fact, we hypothesize that aspirin, in addition to its anti-inflammatory effect, may also specifically inhibit autoimmune response in atherosclerosis by actively increasing CD4+CD25+FOXP3+Treg cells as well as by inducing tolerogenic DCs which induce hyporesponsiveness in responder naïve T cells. If proved to be correct, this hypothesis will provide an opportunity to medical community with an already available aspirin-based immunotherapeutic approach for inducing immune tolerance against atherosclerosis.

Therapeutic cloning by xenotransplanted oocytes, supplemented with species specific reprogramming factors

For therapeutic medication, a lot of work has been done regarding human embryonic stem (ES) cell lines derivation, but immunorejection is a major dilemma of this cell based therapy. Since long time, derivation of patient matched stem cells have been hoped to overcome this problem. Oocytes after nuclear transfer are the most reliable source for patient matched ES cells derivation, for therapeutic use. In humans, utilization of oocytes for stem cell research raises sensitive logistical and ethical questions; primarily surrounding participation of women as oocyte donors. It has been claimed that therapeutic cloning would lead to commercial exploitation of poor women. On the other hand, the therapeutic promise of embryonic stem cell is so huge that there is a strong incentive to find some alternate sources of human oocytes. Here we propose to utilize the cross species oocytes supplemented with human ES cellular extracts to establish patient specific ES cell lines.

Effect of osmotic pressure on spermatozoa characteristics of cryopreserved buffalo bull (Bubalus bubalis) semen

ABSTRACT This study was designed to determine the effect of different osmotic pressures on spermatozoa characteristics of cryopreserved buffalo bull semen using tris egg yolk extender (TEYE). Semen of Nili-Ravi buffalo bulls (n = 4) housed at a semen production unit was collected at weekly intervals for 10 weeks. Three solutions of tris-citric acid-fructose with osmotic pressures of 255, 275 and 295 mOsm/kg were used in extender preparation. Semen straws containing 20 × 106 spermatozoa were processed and stored at −196°C in liquid nitrogen. Post-thaw analyses of spermatozoa included motility, viability, acrosomal integrity, plasma membrane integrity, DNA integrity and lipid per-oxidation. Significantly higher (P < .05) sperm motility, acrosomal and DNA integrity were recorded at osmotic pressures of 275 and 295 mOsm/kg compared to 255 mOsm/kg. However, differences in spermatozoa viability, plasma membrane integrity and lipid per-oxidation were non-significant among three osmotic pressures. It is concluded that osmotic pressure of the solution used in extender preparation plays an important role in post-thaw quality of cryopreserved buffalo bull semen.